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Sun D, Duan X, Li N, Qiao O, Hou Y, Ma Z, Liu S, Gong Y, Liu Z. Construction of ubiquitination-related risk model for predicting prognosis in lung adenocarcinoma. Sci Rep 2025; 15:11787. [PMID: 40189665 PMCID: PMC11973225 DOI: 10.1038/s41598-025-92177-4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2024] [Accepted: 02/25/2025] [Indexed: 04/09/2025] Open
Abstract
Lung adenocarcinoma is the most prevalent lung cancer type. Ubiquitination, a critical post-translational modification process that regulates protein degradation and signaling pathways, has been implicated in various cancers, including LUAD. We aimed to explore the associations between ubiquitination and lung adenocarcinoma. TCGA-LUAD cohort served as the training set. Unsupervised clustering, univariate Cox regression, Random Survival Forests, and least absolute shrinkage and selection operator (LASSO) Cox regression were applied to identify ubiquitination-related genes (URGs), then ubiquitination-related risk scores (URRS) were calculated using gene expression and the univariate Cox's coefficient. Comparisons between the high and the low URRS group regarding chemotherapy drug response, immune infiltration level, tumor mutation burden (TMB), tumor neoantigen load (TNB), PD1/L1 expression, and enriched pathways were performed. URRS was calculated based on the expression of DTL, UBE2S, CISH, and STC1. Patients with higher URRS had a worse prognosis (Hazard Ratio [HR] = 0.54, 95% Confidence Interval [CI]: 0.39-0.73, p < 0.001), and the prognosis of the URRS was further confirmed in 6 external validation cohorts (Hazard Ratio [HR] = 0.58, 95% Confidence Interval [CI]: 0.36-0.93, pmax = 0.023). The high URRS group had higher PD1/L1 expression level (p < 0.05), TMB (p < 0.001), TNB (p < 0.001), and TME scores (p < 0.001). The IC50 values of various chemotherapy drugs were lower in the high URRS group. In addition, we found that upregulation of STC1, UBE2S, and DTL was associated with worse, while upregulation of CISH was associated with better prognosis. We also performed a reverse transcription-quantitative polymerase chain reaction (RT-qPCR) for validation. In conclusion, the ubiquitination-based signature might serve as a biomarker to help evaluate the prognosis, biological features, and appropriate treatment for patients with lung adenocarcinoma.
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Affiliation(s)
- Dawei Sun
- School of Disaster and Emergency Medicine, Faculty of Medicine, Tianjin University, No. 92 Weijin Road, Nankai District, Tianjin, 300072, China
- Institute of Disaster and Emergency Medicine, Faculty of Medicine, Tianjin University, Tianjin, China
- Medical School, Faculty of Medicine, Tianjin University, Tianjin, China
- Beijing ChosenMed Clinical Laboratory Co. Ltd, Beijing, 100176, China
| | - Xiaohong Duan
- School of Disaster and Emergency Medicine, Faculty of Medicine, Tianjin University, No. 92 Weijin Road, Nankai District, Tianjin, 300072, China
- Institute of Disaster and Emergency Medicine, Faculty of Medicine, Tianjin University, Tianjin, China
- Medical School, Faculty of Medicine, Tianjin University, Tianjin, China
| | - Ning Li
- School of Disaster and Emergency Medicine, Faculty of Medicine, Tianjin University, No. 92 Weijin Road, Nankai District, Tianjin, 300072, China
- Institute of Disaster and Emergency Medicine, Faculty of Medicine, Tianjin University, Tianjin, China
- Medical School, Faculty of Medicine, Tianjin University, Tianjin, China
| | - Ou Qiao
- School of Disaster and Emergency Medicine, Faculty of Medicine, Tianjin University, No. 92 Weijin Road, Nankai District, Tianjin, 300072, China
- Institute of Disaster and Emergency Medicine, Faculty of Medicine, Tianjin University, Tianjin, China
- Medical School, Faculty of Medicine, Tianjin University, Tianjin, China
| | - Yingjie Hou
- School of Disaster and Emergency Medicine, Faculty of Medicine, Tianjin University, No. 92 Weijin Road, Nankai District, Tianjin, 300072, China
- Institute of Disaster and Emergency Medicine, Faculty of Medicine, Tianjin University, Tianjin, China
- Medical School, Faculty of Medicine, Tianjin University, Tianjin, China
| | - Zihuan Ma
- Beijing ChosenMed Clinical Laboratory Co. Ltd, Beijing, 100176, China
| | - Siyao Liu
- Beijing ChosenMed Clinical Laboratory Co. Ltd, Beijing, 100176, China
| | - Yanhua Gong
- School of Disaster and Emergency Medicine, Faculty of Medicine, Tianjin University, No. 92 Weijin Road, Nankai District, Tianjin, 300072, China.
- Institute of Disaster and Emergency Medicine, Faculty of Medicine, Tianjin University, Tianjin, China.
- Medical School, Faculty of Medicine, Tianjin University, Tianjin, China.
| | - Zichuan Liu
- School of Pharmaceutical Science and Technology, Faculty of Medicine, Tianjin University, Tianjin, China.
- Frontiers Science Center for Synthetic Biology (Ministry of Education), Tianjin University, Tianjin, China.
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Wang H, Wang Y, Zhou J, Song B, Tu G, Nguyen A, Su J, Coenen F, Wei Z, Rigden DJ, Meng J. Statistical modeling of single-cell epitranscriptomics enabled trajectory and regulatory inference of RNA methylation. CELL GENOMICS 2025; 5:100702. [PMID: 39642887 PMCID: PMC11770222 DOI: 10.1016/j.xgen.2024.100702] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/15/2024] [Revised: 10/07/2024] [Accepted: 11/06/2024] [Indexed: 12/09/2024]
Abstract
As a fundamental mechanism for gene expression regulation, post-transcriptional RNA methylation plays versatile roles in various biological processes and disease mechanisms. Recent advances in single-cell technology have enabled simultaneous profiling of transcriptome-wide RNA methylation in thousands of cells, holding the promise to provide deeper insights into the dynamics, functions, and regulation of RNA methylation. However, it remains a major challenge to determine how to best analyze single-cell epitranscriptomics data. In this study, we developed SigRM, a computational framework for effectively mining single-cell epitranscriptomics datasets with a large cell number, such as those produced by the scDART-seq technique from the SMART-seq2 platform. SigRM not only outperforms state-of-the-art models in RNA methylation site detection on both simulated and real datasets but also provides rigorous quantification metrics of RNA methylation levels. This facilitates various downstream analyses, including trajectory inference and regulatory network reconstruction concerning the dynamics of RNA methylation.
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Affiliation(s)
- Haozhe Wang
- Department of Biosciences and Bioinformatics, Center for Intelligent RNA Therapeutics, Suzhou Key Laboratory of Cancer Biology and Chronic Disease, School of Science, XJTLU Entrepreneur College, Xi'an Jiaotong-Liverpool University, Suzhou, Jiangsu 215123, China; Department of Computer Science, University of Liverpool, L7 8TX Liverpool, UK
| | - Yue Wang
- School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, Jiangsu 210023, China.
| | - Jingxian Zhou
- School of AI and Advanced Computing, XJTLU Entrepreneur College, Xi'an Jiaotong-Liverpool University, Suzhou, Jiangsu 215123, China; Department of Computer Science, University of Liverpool, L7 8TX Liverpool, UK; Sino-French Hoffmann Institute, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou, Guangdong 511436, China
| | - Bowen Song
- Department of Public Health, School of Medicine, Nanjing University of Chinese Medicine, Nanjing, Jiangsu 210023, China; Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China
| | - Gang Tu
- Department of Biosciences and Bioinformatics, Center for Intelligent RNA Therapeutics, Suzhou Key Laboratory of Cancer Biology and Chronic Disease, School of Science, XJTLU Entrepreneur College, Xi'an Jiaotong-Liverpool University, Suzhou, Jiangsu 215123, China
| | - Anh Nguyen
- Department of Computer Science, University of Liverpool, L7 8TX Liverpool, UK
| | - Jionglong Su
- School of AI and Advanced Computing, XJTLU Entrepreneur College, Xi'an Jiaotong-Liverpool University, Suzhou, Jiangsu 215123, China
| | - Frans Coenen
- Department of Computer Science, University of Liverpool, L7 8TX Liverpool, UK
| | - Zhi Wei
- Department of Computer Science, New Jersey Institute of Technology, Newark, NJ 07102, USA
| | - Daniel J Rigden
- Institute of Systems, Molecular and Integrative Biology, University of Liverpool, L7 8TX Liverpool, UK
| | - Jia Meng
- Department of Biosciences and Bioinformatics, Center for Intelligent RNA Therapeutics, Suzhou Key Laboratory of Cancer Biology and Chronic Disease, School of Science, XJTLU Entrepreneur College, Xi'an Jiaotong-Liverpool University, Suzhou, Jiangsu 215123, China; Institute of Biomedical Research, Regulatory Mechanism and Targeted Therapy for Liver Cancer Shiyan Key Laboratory, Hubei Provincial Clinical Research Center for Precise Diagnosis and Treatment of Liver Cancer, Taihe Hospital, Hubei University of Medicine, Shiyan, Hubei 442000, China; Institute of Systems, Molecular and Integrative Biology, University of Liverpool, L7 8TX Liverpool, UK.
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Park JU, Jo JH, Kim S, Redon CE, Aladjem MI, Seo Y, Jang SJ, Jang SM. RepID as a potential biomarker and therapeutic target for lung neuroendocrine tumor. Sci Rep 2024; 14:27487. [PMID: 39523440 PMCID: PMC11551140 DOI: 10.1038/s41598-024-79104-9] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/20/2024] [Accepted: 11/06/2024] [Indexed: 11/16/2024] Open
Abstract
Neuroendocrine tumor (NET) is a rare malignant tumor, notably small cell lung cancer (SCLC), a type of lung neuroendocrine tumor, which has a survival rate of less than 7%. Although various biomarkers including CHGA (Chromogranin A), INSM1 (Insulinoma-associated protein 1), and SYP (Synaptophysin) are extensively used for the diagnostic testing of NET, their diverse specificities and sensitivities are acknowledged as limitations. Here, we demonstrate that RepID (Replication initiation determinant protein), a component of CRL4 (Cullin-RING ubiquitin E3 ligase 4), holds promise as a biomarker for identifying NET and SCLC. Analysis of the Cancer Cell Line Encyclopedia (CCLE) via the CellMinerCDB portal reveals a high correlation between RepID transcript levels and mRNA expression of NE signature genes. Additionally, RepID protein is highly expressed in SCLC patient tissues and a subset of SCLC cell lines. Viability analysis following treatment with pevonedistat and SZL-P1-41 in SCLC cell lines and human SCLC-organoid models indicates that RepID expression determines the sensitivity to CRL-targeting anti-cancer drugs. These findings suggest that RepID represents a novel biomarker for NET and SCLC, and insights from RepID research in these cancers could lead to innovative therapeutic strategies.
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Affiliation(s)
- Jong-Uk Park
- Department of Biochemistry, Chungbuk National University, Cheongju, 28644, Republic of Korea
- Department of Biological Sciences and Biotechnology, Chungbuk National University, Cheongju, 28644, Republic of Korea
| | - Jae-Hyun Jo
- Department of Biochemistry, Chungbuk National University, Cheongju, 28644, Republic of Korea
- Department of Biological Sciences and Biotechnology, Chungbuk National University, Cheongju, 28644, Republic of Korea
| | - Sangjune Kim
- Department of Biological Sciences and Biotechnology, Chungbuk National University, Cheongju, 28644, Republic of Korea
| | - Christophe E Redon
- Developmental Therapeutics Branch, Center for Cancer Research, NCI, NIH, Bethesda, MD, 20892-4255, USA
| | - Mirit I Aladjem
- Developmental Therapeutics Branch, Center for Cancer Research, NCI, NIH, Bethesda, MD, 20892-4255, USA
| | - Yuri Seo
- SG Medical Inc., 3-11, Ogeum-ro 13-gil, Songpa-gu, Seoul, Republic of Korea
| | - Se Jin Jang
- SG Medical Inc., 3-11, Ogeum-ro 13-gil, Songpa-gu, Seoul, Republic of Korea
- Department of Pathology, Asan Medical Center, University of Ulsan Medical College, Seoul, Republic of Korea
| | - Sang-Min Jang
- Department of Biochemistry, Chungbuk National University, Cheongju, 28644, Republic of Korea.
- Department of Biological Sciences and Biotechnology, Chungbuk National University, Cheongju, 28644, Republic of Korea.
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Xuan X, Cao J, Chen L, Zhang J, Qian Y, Huang C. DTL promotes the growth and migration of melanoma cells through the ERK/E2F1/BUB1 axis. Sci Rep 2024; 14:26288. [PMID: 39487277 PMCID: PMC11530538 DOI: 10.1038/s41598-024-76477-9] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/27/2024] [Accepted: 10/14/2024] [Indexed: 11/04/2024] Open
Abstract
Melanoma is the most dangerous form of skin cancer. Hence, a better understanding of molecular mechanisms in melanoma pathogenesis is urgently needed, which provides a new insight into the therapy of melanoma. DTL gene is screened out in melanoma pathogenesis by integrated bioinformatics analysis, and its expression is validated in the tissue and cell samples of melanoma. Forced DTL expression facilitates the proliferation, invasion, migration and EMT of melanoma cells, while DTL knockdown suppresses the biological behavior of melanoma cells. In addition, DTL promotes the malignancy of melanoma in vivo. Mechanistically, BUB1 is the crucial downstream target of DTL. Reduced DTL expression suppresses BUB1 expression, while enhanced DTL expression induces BUB1 upregulation. Rescue experiments showed that growing and migrating of melanoma cells induced by DTL are partially impaired by BUB1 inhibition. In addition, the expression of phosphorylated ERK (p-ERK) and the downstream transcription factor E2F1 are reduced when DTL expression is blocked. Meanwhile, BUB1 levels are decreased when the expression of p-ERK or E2F1 is repressed. Notably, the growth and migration of melanoma cells by inhibition of ERK and knockdown of E2F1 was rescued by overexpressing BUB1. DTL gene may be a prognosis marker and represent a unique potential target for melanoma patients. DTL supports the biologically malignant activity of melanoma cells via the ERK/E2F1/BUB1 axis.
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Affiliation(s)
- Xiuyun Xuan
- Department of Dermatology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, Hubei, China
| | - Juanmei Cao
- Department of Dermatology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, Hubei, China
- Department of Dermatology, The First Affiliated Hospital, Shihezi University, Shihezi, 832061, Xinjiang, China
| | - Li Chen
- Department of Dermatology, Hubei Provincial Hospital of Integrated Chinese and Western Medicine, Wuhan, 430015, Hubei, China
| | - Jing Zhang
- Department of Dermatology, General Hospital of Central Theater Command of Chinese People's Liberation Army, Wuhan, 430070, China.
| | - Yue Qian
- Department of Dermatology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, Hubei, China.
| | - Changzheng Huang
- Department of Dermatology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, Hubei, China.
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Lin Y, Tang W, Huang P, Wang Z, Duan L, Jia C, Sun R, Liu L, Shen J. Denticleless E3 ubiquitin protein ligase (DTL) maintains the proliferation and differentiation of epidermis and hair follicles during skin development. Dev Dyn 2024; 253:635-647. [PMID: 38131461 DOI: 10.1002/dvdy.682] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/27/2023] [Revised: 11/16/2023] [Accepted: 11/20/2023] [Indexed: 12/23/2023] Open
Abstract
BACKGROUND A precise balance between the proliferation and differentiation of epidermal progenitors is required to achieve the barrier function during the development of epidermis. During the entire process of skin development, the newly formed basal layer cells divide, differentiate, and migrate outward to the surface of the skin, which is tightly regulated by a series of events related to cell cycle progression. The CRL4DTL complex (Cullin 4 RING ligase, in association with the substrate receptor DTL) has long emerged as a master regulator in various cellular processes, which mediates the degradation of key cell cycle proteins. However, the roles of DTL in regulating epidermal morphogenesis during skin development remain unclear. RESULTS We showed that DTL deficiency in epidermal progenitor cells leads to defects in epidermal stratification and loss of hair follicles accompanied by reduced epidermal progenitor cells and disturbed cell cycle progression during skin development. Transcriptome analysis revealed that p53 pathway is activated in DTL-depleted epidermal progenitor cells. The apoptosis of epidermal cells showed in DTL deficiency mice is rescued by the absence of p53, but the proliferation and differentiation defects were p53-independent. CONCLUSION Our findings indicate that DTL plays a vital role in epidermal malformation during skin development.
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Affiliation(s)
- Yanhui Lin
- Institute of Life Sciences, College of Life and Environmental Science, Wenzhou University, Wenzhou, China
| | - Weibo Tang
- School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, China
- Laboratory of Tumor Targeted Therapy and Translational Medicine, Jilin Medical University, Jilin, China
| | - Peijun Huang
- School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, China
| | - Zhendong Wang
- Key Laboratory of Interventional Pulmonology of Zhejiang Province, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China
| | - Lian Duan
- Central Laboratory, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China
| | - Chonghui Jia
- Department of Endodontics, The First Affiliated Hospital of Harbin Medical University, Harbin, China
| | - Ruizhen Sun
- Department of Histology and Embryology, Harbin Medical University, Harbin, China
| | - Li Liu
- Institute of Life Sciences, College of Life and Environmental Science, Wenzhou University, Wenzhou, China
| | - Jingling Shen
- Institute of Life Sciences, College of Life and Environmental Science, Wenzhou University, Wenzhou, China
- Department of Histology and Embryology, Harbin Medical University, Harbin, China
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Rivard RS, Chang YC, Ragland RL, Thu YM, Kassab M, Mandal RS, Van Riper SK, Kulej K, Higgins L, Markowski TM, Shang D, Hedberg J, Erber L, Garcia B, Chen Y, Bielinsky AK, Brown EJ. Improved detection of DNA replication fork-associated proteins. Cell Rep 2024; 43:114178. [PMID: 38703364 PMCID: PMC12034227 DOI: 10.1016/j.celrep.2024.114178] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/12/2022] [Revised: 03/06/2024] [Accepted: 04/16/2024] [Indexed: 05/06/2024] Open
Abstract
Innovative methods to retrieve proteins associated with actively replicating DNA have provided a glimpse into the molecular dynamics of replication fork stalling. We report that a combination of density-based replisome enrichment by isolating proteins on nascent DNA (iPOND2) and label-free quantitative mass spectrometry (iPOND2-DRIPPER) substantially increases both replication factor yields and the dynamic range of protein quantification. Replication protein abundance in retrieved nascent DNA is elevated up to 300-fold over post-replicative controls, and recruitment of replication stress factors upon fork stalling is observed at similar levels. The increased sensitivity of iPOND2-DRIPPER permits direct measurement of ubiquitination events without intervening retrieval of diglycine tryptic fragments of ubiquitin. Using this approach, we find that stalled replisomes stimulate the recruitment of a diverse cohort of DNA repair factors, including those associated with poly-K63-ubiquitination. Finally, we uncover the temporally controlled association of stalled replisomes with nuclear pore complex components and nuclear cytoskeleton networks.
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Affiliation(s)
- Rebecca S Rivard
- Department of Cancer Biology and the Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA
| | - Ya-Chu Chang
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN, USA
| | - Ryan L Ragland
- Department of Cancer Biology and the Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA
| | - Yee-Mon Thu
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN, USA
| | - Muzaffer Kassab
- Department of Cancer Biology and the Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA
| | - Rahul Shubhra Mandal
- Department of Cancer Biology and the Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA
| | - Susan K Van Riper
- University of Minnesota Informatics Institute, University of Minnesota, Minneapolis, MN, USA
| | - Katarzyna Kulej
- Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA
| | - LeeAnn Higgins
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN, USA
| | - Todd M Markowski
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN, USA
| | - David Shang
- Department of Cancer Biology and the Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA
| | - Jack Hedberg
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN, USA
| | - Luke Erber
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN, USA
| | - Benjamin Garcia
- Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA
| | - Yue Chen
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN, USA
| | - Anja-Katrin Bielinsky
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN, USA.
| | - Eric J Brown
- Department of Cancer Biology and the Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA.
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Liu D, Wu G, Wang S, Zheng X, Che X. Evaluating the Role of Neddylation Modifications in Kidney Renal Clear Cell Carcinoma: An Integrated Approach Using Bioinformatics, MLN4924 Dosing Experiments, and RNA Sequencing. Pharmaceuticals (Basel) 2024; 17:635. [PMID: 38794205 PMCID: PMC11125012 DOI: 10.3390/ph17050635] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/09/2024] [Revised: 05/07/2024] [Accepted: 05/11/2024] [Indexed: 05/26/2024] Open
Abstract
BACKGROUND Neddylation, a post-translational modification process, plays a crucial role in various human neoplasms. However, its connection with kidney renal clear cell carcinoma (KIRC) remains under-researched. METHODS We validated the Gene Set Cancer Analysis Lite (GSCALite) platform against The Cancer Genome Atlas (TCGA) database, analyzing 33 cancer types and their link with 17 neddylation-related genes. This included examining copy number variations (CNVs), single nucleotide variations (SNVs), mRNA expression, cellular pathway involvement, and methylation. Using Gene Set Variation Analysis (GSVA), we categorized these genes into three clusters and examined their impact on KIRC patient prognosis, drug responses, immune infiltration, and oncogenic pathways. Afterward, our objective is to identify genes that exhibit overexpression in KIRC and are associated with an adverse prognosis. After pinpointing the specific target gene, we used the specific inhibitor MLN4924 to inhibit the neddylation pathway to conduct RNA sequencing and related in vitro experiments to verify and study the specificity and potential mechanisms related to the target. This approach is geared towards enhancing our understanding of the prognostic importance of neddylation modification in KIRC. RESULTS We identified significant CNV, SNV, and methylation events in neddylation-related genes across various cancers, with notably higher expression levels observed in KIRC. Cluster analysis revealed a potential trade-off in the interactions among neddylation-related genes, where both high and low levels of gene expression are linked to adverse prognoses. This association is particularly pronounced concerning lymph node involvement, T stage classification, and Fustat score. Simultaneously, our research discovered that PSMB10 exhibits overexpression in KIRC when compared to normal tissues, negatively impacting patient prognosis. Through RNA sequencing and in vitro assays, we confirmed that the inhibition of neddylation modification could play a role in the regulation of various signaling pathways, thereby influencing the prognosis of KIRC. Moreover, our results underscore PSMB10 as a viable target for therapeutic intervention in KIRC, opening up novel pathways for the development of targeted treatment strategies. CONCLUSION This study underscores the regulatory function and potential mechanism of neddylation modification on the phenotype of KIRC, identifying PSMB10 as a key regulatory target with a significant role in influencing the prognosis of KIRC.
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Affiliation(s)
- Dequan Liu
- Department of Urology, The First Affiliated Hospital of Dalian Medical University, Dalian 116011, China; (D.L.); (G.W.); (S.W.)
| | - Guangzhen Wu
- Department of Urology, The First Affiliated Hospital of Dalian Medical University, Dalian 116011, China; (D.L.); (G.W.); (S.W.)
| | - Shijin Wang
- Department of Urology, The First Affiliated Hospital of Dalian Medical University, Dalian 116011, China; (D.L.); (G.W.); (S.W.)
| | - Xu Zheng
- Department of Cell Biology, College of Basic Medical Science, Dalian Medical University, Dalian 116011, China
| | - Xiangyu Che
- Department of Urology, The First Affiliated Hospital of Dalian Medical University, Dalian 116011, China; (D.L.); (G.W.); (S.W.)
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8
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Tian J, Wen M, Gao P, Feng M, Wei G. RUVBL1 ubiquitination by DTL promotes RUVBL1/2-β-catenin-mediated transcriptional regulation of NHEJ pathway and enhances radiation resistance in breast cancer. Cell Death Dis 2024; 15:259. [PMID: 38609375 PMCID: PMC11015013 DOI: 10.1038/s41419-024-06651-4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/01/2023] [Revised: 02/04/2024] [Accepted: 04/05/2024] [Indexed: 04/14/2024]
Abstract
Radiotherapy effectiveness in breast cancer is limited by radioresistance. Nevertheless, the mechanisms behind radioresistance are not yet fully understood. RUVBL1 and RUVBL2, referred to as RUVBL1/2, are crucial AAA+ ATPases that act as co-chaperones and are connected to cancer. Our research revealed that RUVBL1, also known as pontin/TIP49, is excessively expressed in MMTV-PyMT mouse models undergoing radiotherapy, which is considered a murine spontaneous breast-tumor model. Our findings suggest that RUVBL1 enhances DNA damage repair and radioresistance in breast cancer cells both in vitro and in vivo. Mechanistically, we discovered that DTL, also known as CDT2 or DCAF2, which is a substrate adapter protein of CRL4, promotes the ubiquitination of RUVBL1 and facilitates its binding to RUVBL2 and transcription cofactor β-catenin. This interaction, in turn, attenuates its binding to acetyltransferase Tat-interacting protein 60 (TIP60), a comodulator of nuclear receptors. Subsequently, ubiquitinated RUVBL1 promotes the transcriptional regulation of RUVBL1/2-β-catenin on genes associated with the non-homologous end-joining (NHEJ) repair pathway. This process also attenuates TIP60-mediated H4K16 acetylation and the homologous recombination (HR) repair process. Expanding upon the prior study's discoveries, we exhibited that the ubiquitination of RUVBL1 by DTL advances the interosculation of RUVBL1/2-β-catenin. And, it then regulates the transcription of NHEJ repair pathway protein. Resulting in an elevated resistance of breast cancer cells to radiation therapy. From the aforementioned, it is evident that targeting DTL-RUVBL1/2-β-catenin provides a potential radiosensitization approach when treating breast cancer.
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Affiliation(s)
- Jie Tian
- Key Laboratory for Experimental Teratology of the Ministry of Education, Department of Cell Biology, School of Basic Medical Sciences, Cheeloo College of Medicine, Shandong University, Jinan, Shandong, 250012, China
| | - Mingxin Wen
- Key Laboratory for Experimental Teratology of the Ministry of Education, Department of Human Anatomy, School of Basic Medical Sciences, Cheeloo College of Medicine, Shandong University, Jinan, Shandong, 250012, China
| | - Peng Gao
- Key Laboratory for Experimental Teratology of Ministry of Education, Department of Pathology, School of Basic Medical Sciences and Qilu Hospital, Shandong University, Jinan, Shandong, 250012, China
| | - Maoxiao Feng
- Department of Clinical Laboratory, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, Shandong, China.
| | - Guangwei Wei
- Key Laboratory for Experimental Teratology of the Ministry of Education, Department of Cell Biology, School of Basic Medical Sciences, Cheeloo College of Medicine, Shandong University, Jinan, Shandong, 250012, China.
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9
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Shi J, Yu X, Li G, Zhao X, Chen J, Fang Y, Yang Y, Wang T, Xu T, Bian L, Lyu L, He Y. DTL promotes head and neck squamous cell carcinoma progression by mediating the degradation of ARGLU1 to regulate the Notch signaling pathway. Int J Biol Macromol 2024; 259:129184. [PMID: 38218284 DOI: 10.1016/j.ijbiomac.2023.129184] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/15/2023] [Revised: 12/30/2023] [Accepted: 12/30/2023] [Indexed: 01/15/2024]
Abstract
Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer worldwide, with a high incidence in squamous epithelium. The E3 ubiquitin ligase DTL is a component of the CRL4A complex and is widely involved in tumor progression. We aimed to analyze the role of DTL in HNSCC and to explore its mechanism of action. Through clinical analysis, we found that DTL is upregulated in HNSCC tissues and is associated with the tumor microenvironment and poor survival in patients. Through gain-of-function and loss-of-function assays, we showed that DTL promotes cell proliferation and migration in vitro and tumor growth in vivo. Mass spectrometry analysis and immunoprecipitation assays showed that DTL interacts with ARGLU1 to promote K11-linked ubiquitination-mediated degradation of ARGLU1, thereby promoting the activation of the CSL-dependent Notch signaling pathway. Furthermore, siARGLU1 blocks the inhibitory effects of DTL knockdown on HNSCC cells. In this study, we showed that DTL promotes HNSCC progression through K11-linked ubiquitination of ARGLU1 to activate the CSL-dependent Notch pathway. These findings identify a promising therapeutic target for HNSCC.
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Affiliation(s)
- Jingpei Shi
- Department of Oral and Maxillofacial Surgery, School and Hospital of Stomatology, Yunnan Key Laboratory of Stem Cell and Regenerative Medicine, NHC Key Laboratory of Drug Addiction Medicine, Kunming Medical University, Kunming 650106, Yunnan, China; Yunnan Key Laboratory of Stem Cell and Regenerative Medicine, Science and Technology Achievement Incubation Center, Kunming Medical University, Kunming 650500, Yunnan, China
| | - Xiaonan Yu
- Department of Oral and Maxillofacial Surgery, School and Hospital of Stomatology, Yunnan Key Laboratory of Stem Cell and Regenerative Medicine, NHC Key Laboratory of Drug Addiction Medicine, Kunming Medical University, Kunming 650106, Yunnan, China
| | - Guoyu Li
- Department of Colorectal Surgery, Yunnan Cancer Hospital, The Third Affiliated Hospital of Kunming Medical University, Kunming 650118, Yunnan, China
| | - Xiaoyu Zhao
- Department of Dermatology, The First Affiliated Hospital of Kunming Medical University, Kunming 650032,Yunnan, China
| | - Jiwen Chen
- Yunnan Key Laboratory of Stem Cell and Regenerative Medicine, Science and Technology Achievement Incubation Center, Kunming Medical University, Kunming 650500, Yunnan, China
| | - Ying Fang
- Department of Infection and Hepatology, The First Affiliated Hospital of Kunming Medical University, 650032, Yunnan, China
| | - Yan Yang
- Department of Hepatobiliary and Pancreatic Surgery and Liver Transplantion, the First People's Hospital of Kunming, Kunming 650011, Yunnan, China
| | - Ting Wang
- Yunnan Key Laboratory of Stem Cell and Regenerative Medicine, Science and Technology Achievement Incubation Center, Kunming Medical University, Kunming 650500, Yunnan, China
| | - Tianyong Xu
- Yunnan Key Laboratory of Stem Cell and Regenerative Medicine, Science and Technology Achievement Incubation Center, Kunming Medical University, Kunming 650500, Yunnan, China
| | - Li Bian
- Department of Pathology, The First Affiliated Hospital of Kunming Medical University, Kunming 650032, Yunnan, China.
| | - Lechun Lyu
- Yunnan Key Laboratory of Stem Cell and Regenerative Medicine, Science and Technology Achievement Incubation Center, Kunming Medical University, Kunming 650500, Yunnan, China.
| | - Yongwen He
- Department of Oral and Maxillofacial Surgery, School and Hospital of Stomatology, Yunnan Key Laboratory of Stem Cell and Regenerative Medicine, NHC Key Laboratory of Drug Addiction Medicine, Kunming Medical University, Kunming 650106, Yunnan, China; Qujing Medical College, Qujing 655099, Yunnan, China.
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10
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Zhang Y, Wang C, Wu L, Bai C, Huang K, Yao L, Zhang Z, Ye L, Liu R, Ge X, Xu M, Zhao Y, Cao Q. Epithelial CRL4 DCAF2 Is Critical for Maintaining Intestinal Homeostasis Against DSS-Induced Colitis by Regulating the Proliferation and Repair of Intestinal Epithelial Cells. Dig Dis Sci 2024; 69:66-80. [PMID: 37968554 DOI: 10.1007/s10620-023-08147-1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/19/2023] [Accepted: 06/21/2023] [Indexed: 11/17/2023]
Abstract
BACKGROUND AND AIMS Inflammatory bowel disease (IBD) is currently gaining an increasing global interest. Intestinal epithelial barrier dysfunction is crucial toward developing IBD; however, the underlying mechanisms are not yet elucidated. This study is aimed at elucidating the function of CRL4DCAF2, an E3 ligase, toward mediating intestinal homeostasis. METHODS Colon samples were collected from patients with IBD and healthy individuals to examine the expression of CRL4DCAF2. CRL4DCAF2 conditional knockdown in mouse intestinal epithelial cells (IECs) (DCAF2EKD) were constructed. DCAF2EKD and their littermate control (DCAF2EWT) were treated with dextran sodium sulfate (DSS) to induce acute colitis. Transcriptome analysis was performed on inflamed colon samples obtained from the mice. Cell cycle regulators were evaluated using real-time polymerase chain reaction (PCR), while tight junction and apoptosis proteins were examined via immunofluorescence and western blot. RESULTS CRL4DCAF2 expression was significantly decreased in the inflamed IBD epithelium, and low expression of CRL4DCAF2 associated with high recurrence risk. Mice with DCAF2 specific knockout in IECs suffer from embryonic death. Multiple genes involved in cell proliferation, immune response, and gap junction were differentially expressed in inflamed colon from DCAF2EKD compared with DCAF2EWT. Furthermore, conditional downregulation of CRL4DCAF2 in the intestinal epithelium induced primarily epithelial damage, increased intestinal permeability, and diminished tight junction protein expression. In vivo and in vitro cell transfection experiments revealed that CRL4DCAF2 enhanced cell proliferation by promoting p21 ubiquitination and degradation, thereby inhibiting G2/M cell cycle. In addition, CRL4DCAF2 can also inhibit IEC apoptosis and promote cell autophagy. CONCLUSIONS CRL4DCAF2 downregulation in IECs promotes intestinal barrier dysfunction and inhibits IEC proliferation, thus making it more susceptible to inflammation.
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Affiliation(s)
- Yu Zhang
- Department of Gastroenterology, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Inflammatory Bowel Disease Center, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Institute of Gastroenterology, Zhejiang University, Hangzhou, 310016, Zhejiang, China
| | - Chaohui Wang
- Department of Gastroenterology, Taizhou Central Hospital, Taizhou, 318000, China
| | - Lexi Wu
- Department of Gastroenterology, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Inflammatory Bowel Disease Center, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Institute of Gastroenterology, Zhejiang University, Hangzhou, 310016, Zhejiang, China
| | - Chenhao Bai
- Department of Gastroenterology, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Inflammatory Bowel Disease Center, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Institute of Gastroenterology, Zhejiang University, Hangzhou, 310016, Zhejiang, China
| | - Kaituo Huang
- Department of Gastroenterology, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Inflammatory Bowel Disease Center, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Institute of Gastroenterology, Zhejiang University, Hangzhou, 310016, Zhejiang, China
| | - Lingya Yao
- Department of Gastroenterology, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Inflammatory Bowel Disease Center, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Institute of Gastroenterology, Zhejiang University, Hangzhou, 310016, Zhejiang, China
| | - Zhou Zhang
- Department of Gastroenterology, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Inflammatory Bowel Disease Center, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Institute of Gastroenterology, Zhejiang University, Hangzhou, 310016, Zhejiang, China
| | - Lingna Ye
- Department of Gastroenterology, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Inflammatory Bowel Disease Center, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Institute of Gastroenterology, Zhejiang University, Hangzhou, 310016, Zhejiang, China
| | - Rongbei Liu
- Department of Gastroenterology, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Inflammatory Bowel Disease Center, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Institute of Gastroenterology, Zhejiang University, Hangzhou, 310016, Zhejiang, China
| | - Xiaolong Ge
- Inflammatory Bowel Disease Center, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
| | - Mengque Xu
- Department of Gastroenterology, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Inflammatory Bowel Disease Center, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Institute of Gastroenterology, Zhejiang University, Hangzhou, 310016, Zhejiang, China
| | - Yuan Zhao
- Department of Gastroenterology, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Inflammatory Bowel Disease Center, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
- Institute of Gastroenterology, Zhejiang University, Hangzhou, 310016, Zhejiang, China
| | - Qian Cao
- Department of Gastroenterology, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China.
- Inflammatory Bowel Disease Center, Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China.
- Institute of Gastroenterology, Zhejiang University, Hangzhou, 310016, Zhejiang, China.
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11
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Liu Z, Yang G, Yi X, Zhang S, Feng Z, Cui X, Chen F, Yu L. Osteopontin regulates the growth and invasion of liver cancer cells via DTL. Oncol Lett 2023; 26:476. [PMID: 37809049 PMCID: PMC10551862 DOI: 10.3892/ol.2023.14064] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2023] [Accepted: 07/19/2023] [Indexed: 10/10/2023] Open
Abstract
Osteopontin (OPN), a secreted phosphoglycoprotein, has important roles in tumor growth, invasion and metastasis in numerous types of cancers. Denticleless E3 ubiquitin protein ligase homolog (DTL), one of the CUL4-DDB1-associated factors (DCAFs), has also been associated with the invasion and metastasis of cancer cells. In the present study, OPN was found to induce DTL expression in liver cancer cells, and the results obtained using luciferase activity assays demonstrated that OPN could transcriptionally activate DTL expression in liver cancer cells. Furthermore, the results of the present study demonstrated that OPN could increase the expression of DTL via PI3K/AKT signaling. In conclusion, the present study demonstrated that OPN, as an extracellular matrix protein, is able to promote the growth and invasion of liver cancer cells through stimulation of the expression of DTL via the PI3K/AKT signaling pathway.
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Affiliation(s)
- Zhiyong Liu
- Department of General Interventional Radiology, Guangxi Academy of Medical Sciences and The People's Hospital of Guangxi Zhuang Autonomous Region, Nanning, Guangxi Zhuang Autonomous Region 530021, P.R. China
| | - Guang Yang
- State Key Laboratory of Oncology in South China, Department of Imaging and Interventional Radiology, Cancer Center, Sun Yat-sen University, Guangzhou, Guangdong 510060, P.R. China
| | - Xiaoyu Yi
- Department of General Interventional Radiology, Guangxi Academy of Medical Sciences and The People's Hospital of Guangxi Zhuang Autonomous Region, Nanning, Guangxi Zhuang Autonomous Region 530021, P.R. China
| | - Shijie Zhang
- Department of General Interventional Radiology, Guangxi Academy of Medical Sciences and The People's Hospital of Guangxi Zhuang Autonomous Region, Nanning, Guangxi Zhuang Autonomous Region 530021, P.R. China
| | - Zhibo Feng
- Department of General Interventional Radiology, Guangxi Academy of Medical Sciences and The People's Hospital of Guangxi Zhuang Autonomous Region, Nanning, Guangxi Zhuang Autonomous Region 530021, P.R. China
| | - Xudong Cui
- Department of General Interventional Radiology, Guangxi Academy of Medical Sciences and The People's Hospital of Guangxi Zhuang Autonomous Region, Nanning, Guangxi Zhuang Autonomous Region 530021, P.R. China
| | - Feilong Chen
- Department of General Interventional Radiology, Guangxi Academy of Medical Sciences and The People's Hospital of Guangxi Zhuang Autonomous Region, Nanning, Guangxi Zhuang Autonomous Region 530021, P.R. China
| | - Lei Yu
- Department of General Interventional Radiology, Guangxi Academy of Medical Sciences and The People's Hospital of Guangxi Zhuang Autonomous Region, Nanning, Guangxi Zhuang Autonomous Region 530021, P.R. China
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12
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Singh G, Sharma SK, Dorata A, Singh SK. miR-17 ~ 92 suppresses proliferation and invasion of cervical cancer cells by inhibiting cell cycle regulator Cdt2. Discov Oncol 2023; 14:172. [PMID: 37707654 PMCID: PMC10501107 DOI: 10.1007/s12672-023-00775-3] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/25/2023] [Accepted: 08/21/2023] [Indexed: 09/15/2023] Open
Abstract
Cervical cancer (CC) is the 4th most leading cause of death among women worldwide, and if diagnosed in late stages the treatment options are almost negligible. 99% of CC is caused by high-risk human papilloma viruses (HR-HPV). Upon integration into human genome, the encoded viral proteins mis-regulate various onco-suppressors and checkpoint factors including cell cycle regulators. One such protein is cell cycle S phase licensing factor, CDC-10 dependent transcript-2 (Cdt2) which has been reported to be highly upregulated in various cancers including CC. Also, in CC cells, several tumor suppressor miRNAs are suppressed, including miR-17 ~ 92 cluster. In this study, we report that miR-17 ~ 92 directly recruits to 3'UTR of Cdt2 and downregulates this oncogene which suppresses the proliferation, migration and invasion capabilities of the CC cell lines without affecting non-cancerous cells. We further show that suppression of Cdt2 by miR-17 ~ 92, blocks the cancerous cells in S phase and induces apoptosis, eventually leading to their death. Hence, our work for the first time, mechanistically shows how miR-17 ~ 92 could work as tumor suppressor in cervical cancer cells, opening up the potential of miR-17 ~ 92 to be used in developing therapy for cervical cancer treatment.
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Affiliation(s)
- Garima Singh
- Cell Cycle and Cancer Laboratory, School of Biotechnology, Institute of Science, Banaras Hindu University, Varanasi, UP, 221005, India
| | - Sonika Kumari Sharma
- Cell Cycle and Cancer Laboratory, School of Biotechnology, Institute of Science, Banaras Hindu University, Varanasi, UP, 221005, India
| | - Aastha Dorata
- Cell Cycle and Cancer Laboratory, School of Biotechnology, Institute of Science, Banaras Hindu University, Varanasi, UP, 221005, India
| | - Samarendra Kumar Singh
- Cell Cycle and Cancer Laboratory, School of Biotechnology, Institute of Science, Banaras Hindu University, Varanasi, UP, 221005, India.
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13
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Chen A, Zhou Y, Ren Y, Liu C, Han X, Wang J, Ma Z, Chen Y. Ubiquitination of acetyltransferase Gcn5 contributes to fungal virulence in Fusarium graminearum. mBio 2023; 14:e0149923. [PMID: 37504517 PMCID: PMC10470610 DOI: 10.1128/mbio.01499-23] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/15/2023] [Accepted: 06/16/2023] [Indexed: 07/29/2023] Open
Abstract
The histone acetyltransferase general control non-depressible 5 (Gcn5) plays a critical role in the epigenetic landscape and chromatin modification for regulating a wide variety of biological events. However, the post-translational regulation of Gcn5 itself is poorly understood. Here, we found that Gcn5 was ubiquitinated and deubiquitinated by E3 ligase Tom1 and deubiquitinating enzyme Ubp14, respectively, in the important plant pathogenic fungus Fusarium graminearum. Tom1 interacted with Gcn5 in the nucleus and subsequently ubiquitinated Gcn5 mainly at K252 to accelerate protein degradation. Conversely, Ubp14 deubiquitinated Gcn5 and enhanced its stability. In the deletion mutant Δubp14, protein level of Gcn5 was significantly reduced and resulted in attenuated virulence in the fungus by affecting the mycotoxin production, autophagy process, and the penetration ability. Our findings indicate that Tom1 and Ubp14 show antagonistic functions in the control of the protein stability of Gcn5 via post-translational modification and highlight the importance of Tom1-Gcn5-Ubp14 circuit in the fungal virulence. IMPORTANCE Post-translational modification (PTM) enzymes have been reported to be involved in regulating numerous cellular processes. However, the modification of these PTM enzymes themselves is largely unknown. In this study, we found that the E3 ligase Tom1 and deubiquitinating enzyme Ubp14 contributed to the regulation of ubiquitination and deubiquitination of acetyltransferase Gcn5, respectively, in Fusarium graminearum, the causal agent of Fusarium head blight of cereals. Our findings provide deep insights into the modification of acetyltransferase Gcn5 and its dynamic regulation via ubiquitination and deubiquitination. To our knowledge, this work is the most comprehensive analysis of a regulatory network of ubiquitination that impinges on acetyltransferase in filamentous pathogens. Moreover, our findings are important because we present the novel roles of the Tom1-Gcn5-Ubp14 circuit in fungal virulence, providing novel possibilities and targets to control fungal diseases.
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Affiliation(s)
- Ahai Chen
- State Key Laboratory of Rice Biology and Breeding, Institute of Biotechnology, Zhejiang University, Hangzhou, China
| | - Yifan Zhou
- State Key Laboratory of Rice Biology and Breeding, Institute of Biotechnology, Zhejiang University, Hangzhou, China
| | - Yiyi Ren
- State Key Laboratory of Rice Biology and Breeding, Institute of Biotechnology, Zhejiang University, Hangzhou, China
| | - Chao Liu
- State Key Laboratory of Rice Biology and Breeding, Institute of Biotechnology, Zhejiang University, Hangzhou, China
| | - Xingmin Han
- State Key Laboratory of Rice Biology and Breeding, Institute of Biotechnology, Zhejiang University, Hangzhou, China
| | - Jing Wang
- State Key Laboratory of Rice Biology and Breeding, Institute of Biotechnology, Zhejiang University, Hangzhou, China
| | - Zhonghua Ma
- State Key Laboratory of Rice Biology and Breeding, Institute of Biotechnology, Zhejiang University, Hangzhou, China
| | - Yun Chen
- State Key Laboratory of Rice Biology and Breeding, Institute of Biotechnology, Zhejiang University, Hangzhou, China
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14
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Sinnarasan VSP, Paul D, Das R, Venkatesan A. Gastric Cancer Biomarker Candidates Identified by Machine Learning and Integrative Bioinformatics: Toward Personalized Medicine. OMICS : A JOURNAL OF INTEGRATIVE BIOLOGY 2023. [PMID: 37229622 DOI: 10.1089/omi.2023.0015] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/27/2023]
Abstract
Gastric cancer (GC) is among the leading causes of cancer-related deaths worldwide. The discovery of robust diagnostic biomarkers for GC remains a challenge. This study sought to identify biomarker candidates for GC by integrating machine learning (ML) and bioinformatics approaches. Transcriptome profiles of patients with GC were analyzed to identify differentially expressed genes between the tumor and adjacent normal tissues. Subsequently, we constructed protein-protein interaction networks so as to find the significant hub genes. Along with the bioinformatics integration of ML methods such as support vector machine, the recursive feature elimination was used to select the most informative genes. The analysis unraveled 160 significant genes, with 88 upregulated and 72 downregulated, 10 hub genes, and 12 features from the variable selection method. The integrated analyses found that EXO1, DTL, KIF14, and TRIP13 genes are significant and poised as potential diagnostic biomarkers in relation to GC. The receiver operating characteristic curve analysis found KIF14 and TRIP13 are strongly associated with diagnosis of GC. We suggest KIF14 and TRIP13 are considered as biomarker candidates that might potentially inform future research on diagnosis, prognosis, or therapeutic targets for GC. These findings collectively offer new future possibilities for precision/personalized medicine research and development for patients with GC.
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Affiliation(s)
| | - Dahrii Paul
- Department for Bioinformatics, School of Life Sciences, Pondicherry University, Puducherry, India
| | - Rajesh Das
- Department for Bioinformatics, School of Life Sciences, Pondicherry University, Puducherry, India
| | - Amouda Venkatesan
- Department for Bioinformatics, School of Life Sciences, Pondicherry University, Puducherry, India
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15
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Gioia U, Tavella S, Martínez-Orellana P, Cicio G, Colliva A, Ceccon M, Cabrini M, Henriques AC, Fumagalli V, Paldino A, Presot E, Rajasekharan S, Iacomino N, Pisati F, Matti V, Sepe S, Conte MI, Barozzi S, Lavagnino Z, Carletti T, Volpe MC, Cavalcante P, Iannacone M, Rampazzo C, Bussani R, Tripodo C, Zacchigna S, Marcello A, d'Adda di Fagagna F. SARS-CoV-2 infection induces DNA damage, through CHK1 degradation and impaired 53BP1 recruitment, and cellular senescence. Nat Cell Biol 2023; 25:550-564. [PMID: 36894671 PMCID: PMC10104783 DOI: 10.1038/s41556-023-01096-x] [Citation(s) in RCA: 45] [Impact Index Per Article: 22.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/17/2021] [Accepted: 01/25/2023] [Indexed: 03/11/2023]
Abstract
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the RNA virus responsible for the coronavirus disease 2019 (COVID-19) pandemic. Although SARS-CoV-2 was reported to alter several cellular pathways, its impact on DNA integrity and the mechanisms involved remain unknown. Here we show that SARS-CoV-2 causes DNA damage and elicits an altered DNA damage response. Mechanistically, SARS-CoV-2 proteins ORF6 and NSP13 cause degradation of the DNA damage response kinase CHK1 through proteasome and autophagy, respectively. CHK1 loss leads to deoxynucleoside triphosphate (dNTP) shortage, causing impaired S-phase progression, DNA damage, pro-inflammatory pathways activation and cellular senescence. Supplementation of deoxynucleosides reduces that. Furthermore, SARS-CoV-2 N-protein impairs 53BP1 focal recruitment by interfering with damage-induced long non-coding RNAs, thus reducing DNA repair. Key observations are recapitulated in SARS-CoV-2-infected mice and patients with COVID-19. We propose that SARS-CoV-2, by boosting ribonucleoside triphosphate levels to promote its replication at the expense of dNTPs and by hijacking damage-induced long non-coding RNAs' biology, threatens genome integrity and causes altered DNA damage response activation, induction of inflammation and cellular senescence.
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Affiliation(s)
- Ubaldo Gioia
- IFOM ETS - The AIRC Institute of Molecular Oncology, Milan, Italy
| | - Sara Tavella
- IFOM ETS - The AIRC Institute of Molecular Oncology, Milan, Italy
| | | | - Giada Cicio
- IFOM ETS - The AIRC Institute of Molecular Oncology, Milan, Italy
- University of Palermo, Palermo, Italy
| | - Andrea Colliva
- International Centre for Genetic Engineering and Biotechnology, Trieste, Italy
| | - Marta Ceccon
- IFOM ETS - The AIRC Institute of Molecular Oncology, Milan, Italy
| | - Matteo Cabrini
- IFOM ETS - The AIRC Institute of Molecular Oncology, Milan, Italy
| | - Ana C Henriques
- IFOM ETS - The AIRC Institute of Molecular Oncology, Milan, Italy
| | | | - Alessia Paldino
- International Centre for Genetic Engineering and Biotechnology, Trieste, Italy
- University of Trieste, Trieste, Italy
| | | | - Sreejith Rajasekharan
- International Centre for Genetic Engineering and Biotechnology, Trieste, Italy
- Leibniz Institute for Experimental Virology (HPI), Hamburg, Germany
| | - Nicola Iacomino
- Fondazione IRCCS Istituto Neurologico Carlo Besta, Milan, Italy
| | | | - Valentina Matti
- IFOM ETS - The AIRC Institute of Molecular Oncology, Milan, Italy
| | - Sara Sepe
- IFOM ETS - The AIRC Institute of Molecular Oncology, Milan, Italy
| | - Matilde I Conte
- IFOM ETS - The AIRC Institute of Molecular Oncology, Milan, Italy
| | - Sara Barozzi
- IFOM ETS - The AIRC Institute of Molecular Oncology, Milan, Italy
| | - Zeno Lavagnino
- IFOM ETS - The AIRC Institute of Molecular Oncology, Milan, Italy
| | - Tea Carletti
- International Centre for Genetic Engineering and Biotechnology, Trieste, Italy
| | | | | | - Matteo Iannacone
- IRCCS San Raffaele Scientific Institute & University, Milan, Italy
| | | | | | - Claudio Tripodo
- IFOM ETS - The AIRC Institute of Molecular Oncology, Milan, Italy
- University of Palermo, Palermo, Italy
| | - Serena Zacchigna
- International Centre for Genetic Engineering and Biotechnology, Trieste, Italy
- University of Trieste, Trieste, Italy
| | - Alessandro Marcello
- International Centre for Genetic Engineering and Biotechnology, Trieste, Italy
| | - Fabrizio d'Adda di Fagagna
- IFOM ETS - The AIRC Institute of Molecular Oncology, Milan, Italy.
- Institute of Molecular Genetics (IGM), National Research Institute (CNR), Pavia, Italy.
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16
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Hopkins N. An Editor scientists dream of. Genes Dev 2023; 37:30-31. [PMID: 37061990 PMCID: PMC10046425 DOI: 10.1101/gad.350500.123] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 04/17/2023]
Affiliation(s)
- Nancy Hopkins
- Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA
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17
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Fan Q, Lu Q, Wang G, Zhu W, Teng L, Chen W, Bi L. Optimizing component formula suppresses lung cancer by blocking DTL-mediated PDCD4 ubiquitination to regulate the MAPK/JNK pathway. JOURNAL OF ETHNOPHARMACOLOGY 2022; 299:115546. [PMID: 35850313 DOI: 10.1016/j.jep.2022.115546] [Citation(s) in RCA: 13] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/11/2022] [Revised: 07/02/2022] [Accepted: 07/11/2022] [Indexed: 06/15/2023]
Abstract
ETHNOPHARMACOLOGICAL RELEVANCE Salvia miltiorrhiza Bunge and Panax ginseng C. A. Meyer have special curative effect on cancer treatment. The optimizing component formula (OCF) extracted from those two herbs was in line with the anti-lung cancer treatment principle of activating blood and supplementing 'Qi'. However, the study on the mechanism of component formula has always been an insurmountable challenge. Nowadays, the application of network pharmacology and artificial intelligence (AI) in the field of TCM provides new ideas for the study of new targets and mechanisms of TCM, which promotes the modernization of TCM. AIM OF THE STUDY This study aims to further explore the anti-lung cancer mechanism of OCF by using an integrated strategy of network pharmacology and AI technology. MATERIALS AND METHODS Bioinformatic analysis was used to analyze the expression levels, prognosis and survival of DTL and PDCD4 in cancer patients. The binding strength of OCF and DTL was simulated by molecular docking, and the affinity between them was detected by Bio-layer interferometry. Network pharmacology was used to predict the active components, potential targets and pathways of OCF. The association between key targets and their corresponding components and DTL was analyzed by Ingenuity Pathway Analysis (IPA). MTT assay, colony formation assay, wound-healing assay and transwell assay were used to verify the inhibitory effects of OCF on lung cancer cells in vitro. qRT-PCR and Western blot assay were used to detect the effects of OCF on mRNA and protein expression of DTL, PDCD4 and key genes in MAPK/JNK pathways. RESULTS Bioinformatics analysis showed that DTL was significantly up-regulated in lung cancer, which was associated with high malignancy rate, high metastasis rate and poor prognosis of primary tumor. PDCD4 was down-regulated in lung cancer, and associated with high metastasis rate and poor prognosis. The good affinity between OCF and DTL was predicted and verified by molecular docking and Bio-layer interferometry. Based on the network pharmacological databases, 40 active components and 220 corresponding targets of OCF were screened out. KEGG analysis showed that OCF component targets were mainly enriched in MAPK signaling pathway. IPA results showed the interrelationship between DTL, PDCD4, MAPK pathway genes and their corresponding OCF components. In addition, in vitro experiments demonstrated anti-lung cancer activity of OCF, as validated, via impairing cell viability and cell proliferation, as well as inhibiting migration and invasion abilities in lung cancer cells. qRT-PCR showed that OCF down-regulated the mRNA expression of DTL, MAP4K1, JNK, c-Jun and c-Myc, and up-regulated the mRNA expression of PDCD4 and P53 genes in A549 lung cancer cells. Western blot suggested that OCF suppressed the protein level of DTL and blocked the ubiquitination of PDCD4 in A549 lung cancer cells, and down-regulated the protein levels of MAP4K1, p-JNK and p-c-Jun while up-regulated the proteins expression level of P53. CONCLUSIONS OCF might elicit an anti-lung cancer effect by blocking DTL-mediated PDCD4 ubiquitination and suppression of the MAPK/JNK pathway. Meanwhile, our work revealed that network pharmacology and AI technology strategy are cogent means of studying the active components and mechanism of TCM.
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Affiliation(s)
- Qianqian Fan
- School of Integrated Chinese and Western Medicine, Nanjing University of Chinese Medicine, Nanjing, 210023, China
| | - Qinwei Lu
- School of Integrated Chinese and Western Medicine, Nanjing University of Chinese Medicine, Nanjing, 210023, China
| | - Guiyang Wang
- School of Integrated Chinese and Western Medicine, Nanjing University of Chinese Medicine, Nanjing, 210023, China
| | - Wenjing Zhu
- School of Integrated Chinese and Western Medicine, Nanjing University of Chinese Medicine, Nanjing, 210023, China
| | - Linxin Teng
- School of Integrated Chinese and Western Medicine, Nanjing University of Chinese Medicine, Nanjing, 210023, China
| | - Weiping Chen
- School of Integrated Chinese and Western Medicine, Nanjing University of Chinese Medicine, Nanjing, 210023, China
| | - Lei Bi
- School of Integrated Chinese and Western Medicine, Nanjing University of Chinese Medicine, Nanjing, 210023, China.
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Tu J, Yu S, Li J, Ren M, Zhang Y, Luo J, Sun K, Lv Y, Han Y, Huang Y, Ren X, Jiang T, Tang Z, Williams MTS, Lu Q, Liu M. Dhx38 is required for the maintenance and differentiation of erythro-myeloid progenitors and hematopoietic stem cells by alternative splicing. Development 2022; 149:276218. [DOI: 10.1242/dev.200450] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/10/2021] [Accepted: 07/21/2022] [Indexed: 11/20/2022]
Abstract
ABSTRACT
Mutations that occur in RNA-splicing machinery may contribute to hematopoiesis-related diseases. How splicing factor mutations perturb hematopoiesis, especially in the differentiation of erythro-myeloid progenitors (EMPs), remains elusive. Dhx38 is a pre-mRNA splicing-related DEAH box RNA helicase, for which the physiological functions and splicing mechanisms during hematopoiesis currently remain unclear. Here, we report that Dhx38 exerts a broad effect on definitive EMPs as well as the differentiation and maintenance of hematopoietic stem and progenitor cells (HSPCs). In dhx38 knockout zebrafish, EMPs and HSPCs were found to be arrested in mitotic prometaphase, accompanied by a ‘grape’ karyotype, owing to the defects in chromosome alignment. Abnormal alternatively spliced genes related to chromosome segregation, the microtubule cytoskeleton, cell cycle kinases and DNA damage were present in the dhx38 mutants. Subsequently, EMPs and HSPCs in dhx38 mutants underwent P53-dependent apoptosis. This study provides novel insights into alternative splicing regulated by Dhx38, a process that plays a crucial role in the proliferation and differentiation of fetal EMPs and HSPCs.
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Affiliation(s)
- Jiayi Tu
- Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology 1 , Wuhan 430074 , P.R. China
| | - Shanshan Yu
- Institute of Visual Neuroscience and Stem Cell Engineering, College of Life Sciences and Health, Wuhan University of Science and Technology 2 , Wuhan, Hubei 430065 , P.R. China
| | - Jingzhen Li
- Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology 1 , Wuhan 430074 , P.R. China
| | - Mengmeng Ren
- Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology 1 , Wuhan 430074 , P.R. China
| | - Yangjun Zhang
- Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology 3 , Wuhan 430030 , P.R. China
| | - Jiong Luo
- Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology 1 , Wuhan 430074 , P.R. China
| | - Kui Sun
- Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology 1 , Wuhan 430074 , P.R. China
| | - Yuexia Lv
- Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology 1 , Wuhan 430074 , P.R. China
| | - Yunqiao Han
- Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology 1 , Wuhan 430074 , P.R. China
| | - Yuwen Huang
- Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology 1 , Wuhan 430074 , P.R. China
| | - Xiang Ren
- Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology 1 , Wuhan 430074 , P.R. China
| | - Tao Jiang
- Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology 1 , Wuhan 430074 , P.R. China
| | - Zhaohui Tang
- Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology 1 , Wuhan 430074 , P.R. China
| | - Mark Thomas Shaw Williams
- Charles Oakley Laboratories 4 , Department of Biological and Biomedical Sciences , , Glasgow G4 0BA , UK
- Glasgow Caledonian University 4 , Department of Biological and Biomedical Sciences , , Glasgow G4 0BA , UK
| | - Qunwei Lu
- Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology 1 , Wuhan 430074 , P.R. China
| | - Mugen Liu
- Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology 1 , Wuhan 430074 , P.R. China
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19
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Mice lacking DCAF2 in placenta die at the gastrulation stage. Cell Tissue Res 2022; 389:559-572. [PMID: 35711069 DOI: 10.1007/s00441-022-03655-4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/06/2021] [Accepted: 06/10/2022] [Indexed: 11/02/2022]
Abstract
UV-damaged DNA-binding protein 1 (DDB1) and cullin 4-associated factor 2 (DCAF2, also known as DTL or CDT2) is an evolutionarily highly conserved substrate recognition factor in the cullin 4 RING E3 ubiquitin ligase (CRL4) complex. This complex degrades multiple DNA replication and cell cycle-associated proteins to maintain genome stability. To clarify the function of DCAF2 in vivo, we used Cre recombinase driven by the Elf5 promoter to generate knockout mouse model that was specifically deleted Dcaf2 in the trophoblast lineage (Elf5-Cre; Dcaf2fl/fl, Dcaf2 cKO). Here, we show that mice with the genotype Elf5-Cre; Dcaf2fl/+ are normal and fertile. However, after mating of Elf5-Cre; Dcaf2fl/+ mice with Dcaf2fl/fl, no Dcaf2 cKO pups were born. Timed pregnancy studies have shown that Dcaf2 cKO mice developed abnormally on embryonic day 5.5 and died at gastrulation stage. It is worth noting that the extraembryonic ectoderm of Dcaf2 cKO mice is severely reduced or missing and leading to embryonic death. We also proved that stronger DNA damage accumulated in the trophoblastic cells of Dcaf2 cKO mice at E8.5. In addition, higher expression of Caspase-3 was found in the embryonic and trophoblastic cells of these cKO mice. In general, our research shows that the placental DCAF2 is crucial to the formation of gastrula.
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20
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CRL4Cdt2 Ubiquitin Ligase, A Genome Caretaker Controlled by Cdt2 Binding to PCNA and DNA. Genes (Basel) 2022; 13:genes13020266. [PMID: 35205311 PMCID: PMC8871960 DOI: 10.3390/genes13020266] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2021] [Revised: 01/25/2022] [Accepted: 01/27/2022] [Indexed: 12/22/2022] Open
Abstract
The ubiquitin ligase CRL4Cdt2 plays a vital role in preserving genomic integrity by regulating essential proteins during S phase and after DNA damage. Deregulation of CRL4Cdt2 during the cell cycle can cause DNA re-replication, which correlates with malignant transformation and tumor growth. CRL4Cdt2 regulates a broad spectrum of cell cycle substrates for ubiquitination and proteolysis, including Cdc10-dependent transcript 1 or Chromatin licensing and DNA replication factor 1 (Cdt1), histone H4K20 mono-methyltransferase (Set8) and cyclin-dependent kinase inhibitor 1 (p21), which regulate DNA replication. However, the mechanism it operates via its substrate receptor, Cdc10-dependent transcript 2 (Cdt2), is not fully understood. This review describes the essential features of the N-terminal and C-terminal parts of Cdt2 that regulate CRL4 ubiquitination activity, including the substrate recognition domain, intrinsically disordered region (IDR), phosphorylation sites, the PCNA-interacting protein-box (PIP) box motif and the DNA binding domain. Drugs targeting these specific domains of Cdt2 could have potential for the treatment of cancer.
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21
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DTL Is a Prognostic Biomarker and Promotes Bladder Cancer Progression through Regulating the AKT/mTOR axis. OXIDATIVE MEDICINE AND CELLULAR LONGEVITY 2022; 2022:3369858. [PMID: 35103094 PMCID: PMC8799954 DOI: 10.1155/2022/3369858] [Citation(s) in RCA: 15] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 09/10/2021] [Accepted: 12/23/2021] [Indexed: 01/10/2023]
Abstract
Background Denticleless E3 ubiquitin protein ligase homolog (DTL) has been reported to be an important regulator for tumorigenesis and progression. Nonetheless, the biological functions and molecular mechanisms of DTL in BCa remain elusive. Methods We implemented integrative bioinformatics analysis to explore the diagnostic and prognostic values of DTL based on The Cancer Genome Atlas (TCGA), ArrayExpress, and Gene Expression Omnibus (GEO) databases. Then, we utilized qRT-PCR and immunohistochemistry to verify the clinical significance of DTL expression according to clinical specimens and tissue microarray (TMA). Moreover, the biological functions and underlying mechanisms of DTL in BCa were investigated through in vitro and in vivo experiments. Results Integrative bioinformatics analysis revealed that DTL was a key gene associated with BCa progression, and increased DTL expression was correlated with malignant biological behavior and poor prognosis. Experiments on clinical specimens and tissue microarray (TMA) further confirmed our findings. Bioinformatics analysis demonstrated that DTL could be associated with cell cycle- and DNA replication-associated pathways in BCa. The suppression of DTL inhibited BCa cell proliferation, migration, and invasion in vivo and in vitro. Mechanistically, DTL may promote BCa progression through the AKT/mTOR pathway. Conclusions Increased DTL expression was correlated with malignant biological behavior and poor prognosis of BCa patients, and it may promote BCa progression through the AKT/mTOR pathway. Our research provided a potential predictor and therapeutic target for BCa.
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22
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Wittig KA, Sansam CG, Noble TD, Goins D, Sansam CL. The CRL4DTL E3 ligase induces degradation of the DNA replication initiation factor TICRR/TRESLIN specifically during S phase. Nucleic Acids Res 2021; 49:10507-10523. [PMID: 34534348 PMCID: PMC8501952 DOI: 10.1093/nar/gkab805] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/19/2021] [Revised: 08/26/2021] [Accepted: 09/13/2021] [Indexed: 01/02/2023] Open
Abstract
A DNA replication program, which ensures that the genome is accurately and wholly replicated, is established during G1, before the onset of S phase. In G1, replication origins are licensed, and upon S phase entry, a subset of these will form active replisomes. Tight regulation of the number of active replisomes is crucial to prevent replication stress-induced DNA damage. TICRR/TRESLIN is essential for DNA replication initiation, and the level of TICRR and its phosphorylation determine the number of origins that initiate during S phase. However, the mechanisms regulating TICRR protein levels are unknown. Therefore, we set out to define the TICRR/TRESLIN protein dynamics throughout the cell cycle. Here, we show that TICRR levels are high during G1 and dramatically decrease as cells enter S phase and begin DNA replication. We show that degradation of TICRR occurs specifically during S phase and depends on ubiquitin ligases and proteasomal degradation. Using two targeted siRNA screens, we identify CRL4DTL as a cullin complex necessary for TICRR degradation. We propose that this mechanism moderates the level of TICRR protein available for replication initiation, ensuring the proper number of active origins as cells progress through S phase.
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Affiliation(s)
- Kimberlie A Wittig
- Department of Cell Biology, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA.,Cell Cycle and Cancer Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK 73104, USA
| | - Courtney G Sansam
- Cell Cycle and Cancer Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK 73104, USA
| | - Tyler D Noble
- Department of Cell Biology, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA.,Cell Cycle and Cancer Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK 73104, USA
| | - Duane Goins
- Cell Cycle and Cancer Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK 73104, USA
| | - Christopher L Sansam
- Department of Cell Biology, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA.,Cell Cycle and Cancer Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK 73104, USA
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23
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Wu K, Hopkins BD, Sanchez R, DeVita RJ, Pan ZQ. Targeting Cullin-RING E3 Ubiquitin Ligase 4 by Small Molecule Modulators. JOURNAL OF CELLULAR SIGNALING 2021; 2:195-205. [PMID: 34604860 PMCID: PMC8486283 DOI: 10.33696/signaling.2.051] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 11/27/2022]
Abstract
Cullin-RING E3 ubiquitin ligase 4 (CRL4) plays an essential role in cell cycle progression. Recent efforts using high throughput screening and follow up hit-to-lead studies have led to identification of small molecules 33-11 and KH-4-43 that inhibit E3 CRL4's core ligase complex and exhibit anticancer potential. This review provides: 1) an updated perspective of E3 CRL4, including structural organization, major substrate targets and role in cancer; 2) a discussion of the challenges and strategies for finding the CRL inhibitor; and 3) a summary of the properties of the identified CRL4 inhibitors as well as a perspective on their potential utility to probe CRL4 biology and act as therapeutic agents.
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Affiliation(s)
- Kenneth Wu
- Department of Oncological Sciences, The Icahn School of Medicine at Mount Sinai, One Gustave L. Levy Place, New York, NY 10029-6574, USA
| | - Benjamin D Hopkins
- Department of Oncological Sciences, The Icahn School of Medicine at Mount Sinai, One Gustave L. Levy Place, New York, NY 10029-6574, USA.,Genetics and Genomics, The Icahn School of Medicine at Mount Sinai, One Gustave L. Levy Place, New York, NY 10029-6574, USA
| | - Roberto Sanchez
- Department of Pharmacological Sciences, The Icahn School of Medicine at Mount Sinai, One Gustave L. Levy Place, New York, NY 10029-6574, USA.,Drug Discovery Institute, The Icahn School of Medicine at Mount Sinai, One Gustave L. Levy Place, New York, NY 10029-6574, USA
| | - Robert J DeVita
- Department of Pharmacological Sciences, The Icahn School of Medicine at Mount Sinai, One Gustave L. Levy Place, New York, NY 10029-6574, USA.,Drug Discovery Institute, The Icahn School of Medicine at Mount Sinai, One Gustave L. Levy Place, New York, NY 10029-6574, USA
| | - Zhen-Qiang Pan
- Department of Oncological Sciences, The Icahn School of Medicine at Mount Sinai, One Gustave L. Levy Place, New York, NY 10029-6574, USA
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24
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Coordinating DNA Replication and Mitosis through Ubiquitin/SUMO and CDK1. Int J Mol Sci 2021; 22:ijms22168796. [PMID: 34445496 PMCID: PMC8395760 DOI: 10.3390/ijms22168796] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/20/2021] [Revised: 08/11/2021] [Accepted: 08/12/2021] [Indexed: 12/30/2022] Open
Abstract
Post-translational modification of the DNA replication machinery by ubiquitin and SUMO plays key roles in the faithful duplication of the genetic information. Among other functions, ubiquitination and SUMOylation serve as signals for the extraction of factors from chromatin by the AAA ATPase VCP. In addition to the regulation of DNA replication initiation and elongation, we now know that ubiquitination mediates the disassembly of the replisome after DNA replication termination, a process that is essential to preserve genomic stability. Here, we review the recent evidence showing how active DNA replication restricts replisome ubiquitination to prevent the premature disassembly of the DNA replication machinery. Ubiquitination also mediates the removal of the replisome to allow DNA repair. Further, we discuss the interplay between ubiquitin-mediated replisome disassembly and the activation of CDK1 that is required to set up the transition from the S phase to mitosis. We propose the existence of a ubiquitin–CDK1 relay, where the disassembly of terminated replisomes increases CDK1 activity that, in turn, favors the ubiquitination and disassembly of more replisomes. This model has important implications for the mechanism of action of cancer therapies that induce the untimely activation of CDK1, thereby triggering premature replisome disassembly and DNA damage.
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25
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Khalafi M, Reza Soltani A, Golalipour M, Azimmohseni M, Najafiamiri F. On the spectral coherence between two periodically correlated processes. CAN J STAT 2021. [DOI: 10.1002/cjs.11632] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/05/2022]
Affiliation(s)
- Mahnaz Khalafi
- Department of Statistics, Faculty of Science Golestan University Gorgan Iran
| | - Ahmad Reza Soltani
- Department of Statistics and Operations Research, College of Science Kuwait University Safat Kuwait
| | - Masoud Golalipour
- Medical Cellular and Molecular Research Center Golestan University of Medical Sciences Gorgan Iran
| | - Majid Azimmohseni
- Department of Statistics, Faculty of Science Golestan University Gorgan Iran
| | - Farzad Najafiamiri
- Department of Statistics, Faculty of Science Golestan University Gorgan Iran
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26
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Zhang H. Regulation of DNA Replication Licensing and Re-Replication by Cdt1. Int J Mol Sci 2021; 22:ijms22105195. [PMID: 34068957 PMCID: PMC8155957 DOI: 10.3390/ijms22105195] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/20/2021] [Revised: 05/10/2021] [Accepted: 05/12/2021] [Indexed: 11/29/2022] Open
Abstract
In eukaryotic cells, DNA replication licensing is precisely regulated to ensure that the initiation of genomic DNA replication in S phase occurs once and only once for each mitotic cell division. A key regulatory mechanism by which DNA re-replication is suppressed is the S phase-dependent proteolysis of Cdt1, an essential replication protein for licensing DNA replication origins by loading the Mcm2-7 replication helicase for DNA duplication in S phase. Cdt1 degradation is mediated by CRL4Cdt2 ubiquitin E3 ligase, which further requires Cdt1 binding to proliferating cell nuclear antigen (PCNA) through a PIP box domain in Cdt1 during DNA synthesis. Recent studies found that Cdt2, the specific subunit of CRL4Cdt2 ubiquitin E3 ligase that targets Cdt1 for degradation, also contains an evolutionarily conserved PIP box-like domain that mediates the interaction with PCNA. These findings suggest that the initiation and elongation of DNA replication or DNA damage-induced repair synthesis provide a novel mechanism by which Cdt1 and CRL4Cdt2 are both recruited onto the trimeric PCNA clamp encircling the replicating DNA strands to promote the interaction between Cdt1 and CRL4Cdt2. The proximity of PCNA-bound Cdt1 to CRL4Cdt2 facilitates the destruction of Cdt1 in response to DNA damage or after DNA replication initiation to prevent DNA re-replication in the cell cycle. CRL4Cdt2 ubiquitin E3 ligase may also regulate the degradation of other PIP box-containing proteins, such as CDK inhibitor p21 and histone methylase Set8, to regulate DNA replication licensing, cell cycle progression, DNA repair, and genome stability by directly interacting with PCNA during DNA replication and repair synthesis.
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Affiliation(s)
- Hui Zhang
- Department of Chemistry and Biochemistry, Nevada Institute of Personalized Medicine, University of Nevada, Las Vegas, 4505 South Maryland Parkway, Box 454003, Las Vegas, NV 89154, USA
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27
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Qu Y, Chen C, Chen X, Hao Y, She H, Wang M, Ericson PGP, Lin H, Cai T, Song G, Jia C, Chen C, Zhang H, Li J, Liang L, Wu T, Zhao J, Gao Q, Zhang G, Zhai W, Zhang C, Zhang YE, Lei F. The evolution of ancestral and species-specific adaptations in snowfinches at the Qinghai-Tibet Plateau. Proc Natl Acad Sci U S A 2021; 118:e2012398118. [PMID: 33753478 PMCID: PMC8020664 DOI: 10.1073/pnas.2012398118] [Citation(s) in RCA: 16] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
Species in a shared environment tend to evolve similar adaptations under the influence of their phylogenetic context. Using snowfinches, a monophyletic group of passerine birds (Passeridae), we study the relative roles of ancestral and species-specific adaptations to an extreme high-elevation environment, the Qinghai-Tibet Plateau. Our ancestral trait reconstruction shows that the ancestral snowfinch occupied high elevations and had a larger body mass than most nonsnowfinches in Passeridae. Subsequently, this phenotypic adaptation diversified in the descendant species. By comparing high-quality genomes from representatives of the three phylogenetic lineages, we find that about 95% of genes under positive selection in the descendant species are different from those in the ancestor. Consistently, the biological functions enriched for these species differ from those of their ancestor to various degrees (semantic similarity values ranging from 0.27 to 0.5), suggesting that the three descendant species have evolved divergently from the initial adaptation in their common ancestor. Using a functional assay to a highly selective gene, DTL, we demonstrate that the nonsynonymous substitutions in the ancestor and descendant species have improved the repair capacity of ultraviolet-induced DNA damage. The repair kinetics of the DTL gene shows a twofold to fourfold variation across the ancestor and the descendants. Collectively, this study reveals an exceptional case of adaptive evolution to high-elevation environments, an evolutionary process with an initial adaptation in the common ancestor followed by adaptive diversification of the descendant species.
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Affiliation(s)
- Yanhua Qu
- Key Laboratory of Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, 100101 Beijing, China;
| | - Chunhai Chen
- BGI Genomics, BGI-Shenzhen, 518084 Shenzhen, China
| | - Xiumin Chen
- Key Laboratory of Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, 100101 Beijing, China
| | - Yan Hao
- Key Laboratory of Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, 100101 Beijing, China
- College of Life Science, University of Chinese Academy of Sciences, 100049 Beijing, China
| | - Huishang She
- Key Laboratory of Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, 100101 Beijing, China
- College of Life Science, University of Chinese Academy of Sciences, 100049 Beijing, China
| | - Mengxia Wang
- Key Laboratory of Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, 100101 Beijing, China
- College of Life Science, University of Chinese Academy of Sciences, 100049 Beijing, China
| | - Per G P Ericson
- Department of Bioinformatics and Genetics, Swedish Museum of Natural History, SE-104 05 Stockholm, Sweden
| | - Haiyan Lin
- Key Laboratory of Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, 100101 Beijing, China
| | - Tianlong Cai
- Key Laboratory of Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, 100101 Beijing, China
| | - Gang Song
- Key Laboratory of Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, 100101 Beijing, China
| | - Chenxi Jia
- Key Laboratory of Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, 100101 Beijing, China
| | - Chunyan Chen
- Key Laboratory of Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, 100101 Beijing, China
| | - Hailin Zhang
- BGI Genomics, BGI-Shenzhen, 518084 Shenzhen, China
| | - Jiang Li
- BGI Genomics, BGI-Shenzhen, 518084 Shenzhen, China
| | - Liping Liang
- BGI Genomics, BGI-Shenzhen, 518084 Shenzhen, China
| | - Tianyu Wu
- BGI Genomics, BGI-Shenzhen, 518084 Shenzhen, China
| | - Jinyang Zhao
- BGI Genomics, BGI-Shenzhen, 518084 Shenzhen, China
| | - Qiang Gao
- BGI Genomics, BGI-Shenzhen, 518084 Shenzhen, China
| | - Guojie Zhang
- BGI-Shenzhen, 518083 Shenzhen, China
- State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, 650223 Kunming, China
- Center for Excellence in Animal Evolution and Genetics, Chinese Academy of Sciences, 650223 Kunming, China
- Section for Ecology and Evolution, Department of Biology, University of Copenhagen, DK-2100 Copenhagen, Denmark
| | - Weiwei Zhai
- Key Laboratory of Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, 100101 Beijing, China
- Center for Excellence in Animal Evolution and Genetics, Chinese Academy of Sciences, 650223 Kunming, China
| | - Chi Zhang
- BGI Genomics, BGI-Shenzhen, 518084 Shenzhen, China;
| | - Yong E Zhang
- Key Laboratory of Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, 100101 Beijing, China;
- College of Life Science, University of Chinese Academy of Sciences, 100049 Beijing, China
- Center for Excellence in Animal Evolution and Genetics, Chinese Academy of Sciences, 650223 Kunming, China
- Chinese Institute for Brain Research, 102206 Beijing, China
| | - Fumin Lei
- Key Laboratory of Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, 100101 Beijing, China;
- College of Life Science, University of Chinese Academy of Sciences, 100049 Beijing, China
- Center for Excellence in Animal Evolution and Genetics, Chinese Academy of Sciences, 650223 Kunming, China
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CRL4Cdt2 ubiquitin ligase regulates Dna2 and Rad16 (XPF) nucleases by targeting Pxd1 for degradation. PLoS Genet 2020; 16:e1008933. [PMID: 32692737 PMCID: PMC7394458 DOI: 10.1371/journal.pgen.1008933] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/08/2020] [Revised: 07/31/2020] [Accepted: 06/15/2020] [Indexed: 01/19/2023] Open
Abstract
Structure-specific endonucleases (SSEs) play key roles in DNA replication, recombination, and repair. SSEs must be tightly regulated to ensure genome stability but their regulatory mechanisms remain incompletely understood. Here, we show that in the fission yeast Schizosaccharomyces pombe, the activities of two SSEs, Dna2 and Rad16 (ortholog of human XPF), are temporally controlled during the cell cycle by the CRL4Cdt2 ubiquitin ligase. CRL4Cdt2 targets Pxd1, an inhibitor of Dna2 and an activator of Rad16, for degradation in S phase. The ubiquitination and degradation of Pxd1 is dependent on CRL4Cdt2, PCNA, and a PCNA-binding degron motif on Pxd1. CRL4Cdt2-mediated Pxd1 degradation prevents Pxd1 from interfering with the normal S-phase functions of Dna2. Moreover, Pxd1 degradation leads to a reduction of Rad16 nuclease activity in S phase, and restrains Rad16-mediated single-strand annealing, a hazardous pathway of repairing double-strand breaks. These results demonstrate a new role of the CRL4Cdt2 ubiquitin ligase in genome stability maintenance and shed new light on how SSE activities are regulated during the cell cycle. Structure-specific endonucleases are enzymes that process DNA intermediates generated in DNA replication, recombination, and repair. Proper regulation of these enzymes is critical for maintaining genome stability. Dna2 and XPF are two such enzymes present across eukaryotes, from yeasts to humans. Here, we show that in the fission yeast Schizosaccharomyces pombe, the activities of Dna2 and Rad16 (the equivalent of human XPF) are temporally controlled during the cell cycle by the CRL4Cdt2 ubiquitin E3 ligase. In the S phase of the cell cycle, CRL4Cdt2 promotes the degradation of Pxd1, which is an inhibitor of Dna2 and an activator of Rad16. Through targeting Pxd1 for degradation, CRL4Cdt2 increases the activity of Dna2 in S phase and is important for the normal S-phase function of Dna2. Meanwhile, the degradation of Pxd1 reduces the activity of Rad16 in S phase, and curtails Rad16-dependent single-strand annealing, a mutagenic DNA repair pathway. Our findings uncover a new mechanism regulating two important endonucleases during the cell cycle, and reveal a new way of coordinating endonucleases to safeguard genome stability.
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Xue JM, Liu Y, Wan LH, Zhu YX. Comprehensive Analysis of Differential Gene Expression to Identify Common Gene Signatures in Multiple Cancers. Med Sci Monit 2020; 26:e919953. [PMID: 32035007 PMCID: PMC7027371 DOI: 10.12659/msm.919953] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
Background With the development of research on cancer genomics and microenvironment, a new era of oncology focusing on the complicated gene regulation of pan-cancer research and cancer immunotherapy is emerging. This study aimed to identify the common gene expression characteristics of multiple cancers – lung cancer, liver cancer, kidney cancer, cervical cancer, and breast cancer – and the potential therapeutic targets in public databases. Material/Methods Gene expression analysis of GSE42568, GSE19188, GSE121248, GSE63514, and GSE66272 in the GEO database of multitype cancers revealed differentially expressed genes (DEGs). Then, GO analysis, KEGG function, and path enrichment analyses were performed. Hub-genes were identified by using the degree of association of protein interaction networks. Moreover, the expression of hub-genes in cancers was verified, and hub-gene-related survival analysis was conducted. Finally, infiltration levels of tumor immune cells with related genes were explored. Results We found 12 cross DEGs in the 5 databases (screening conditions: “adj p<0.05” and “logFC>2 or logFC<–2”). The biological processes of DEGs were mainly concentrated in cell division, regulation of chromosome segregation, nuclear division, cell cycle checkpoint, and mitotic nuclear division. Furthermore, 10 hub-genes were obtained using Cytoscape: TOP2A, ECT2, RRM2, ANLN, NEK2, ASPM, BUB1B, CDK1, DTL, and PRC1. The high expression levels of the 10 genes were associated with the poor survival of these multiple cancers, as well as ASPM, may be associated with immune cell infiltration. Conclusions Analysis of the common DEGs of multiple cancers showed that 10 hub-genes, especially ASPM and CDK1, can become potential therapeutic targets. This study can serve as a reference to understand the characteristics of different cancers, design basket clinical trials, and create personalized treatments.
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Affiliation(s)
- Jin-Min Xue
- Department of Oncology, The First Affiliated Hospital of Chongqing Medical University, Chongqing, China (mainland).,Department of Oncology, Jinshan Hospital of The First Affiliated Hospital of Chongqing Medical University, Chongqing, China (mainland).,Chongqing Clinical Cancer Research Center, Chongqing, China (mainland)
| | - Yi Liu
- Department of Oncology, The First Affiliated Hospital of Chongqing Medical University, Chongqing, China (mainland).,Department of Oncology, Jinshan Hospital of The First Affiliated Hospital of Chongqing Medical University, Chongqing, China (mainland).,Chongqing Clinical Cancer Research Center, Chongqing, China (mainland)
| | - Ling-Hong Wan
- Department of Oncology, The First Affiliated Hospital of Chongqing Medical University, Chongqing, China (mainland).,Department of Oncology, Jinshan Hospital of The First Affiliated Hospital of Chongqing Medical University, Chongqing, China (mainland).,Chongqing Clinical Cancer Research Center, Chongqing, China (mainland)
| | - Yu-Xi Zhu
- Department of Oncology, The First Affiliated Hospital of Chongqing Medical University, Chongqing, China (mainland).,Department of Oncology, Jinshan Hospital of The First Affiliated Hospital of Chongqing Medical University, Chongqing, China (mainland).,Chongqing Clinical Cancer Research Center, Chongqing, China (mainland)
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Jang SM, Nathans JF, Fu H, Redon CE, Jenkins LM, Thakur BL, Pongor LS, Baris AM, Gross JM, OʹNeill MJ, Indig FE, Cappell SD, Aladjem MI. The RepID-CRL4 ubiquitin ligase complex regulates metaphase to anaphase transition via BUB3 degradation. Nat Commun 2020; 11:24. [PMID: 31911655 PMCID: PMC6946706 DOI: 10.1038/s41467-019-13808-9] [Citation(s) in RCA: 22] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/04/2019] [Accepted: 11/18/2019] [Indexed: 12/14/2022] Open
Abstract
The spindle assembly checkpoint (SAC) prevents premature chromosome segregation by inactivating the anaphase promoting complex/cyclosome (APC/C) until all chromosomes are properly attached to mitotic spindles. Here we identify a role for Cullin–RING ubiquitin ligase complex 4 (CRL4), known for modulating DNA replication, as a crucial mitotic regulator that triggers the termination of the SAC and enables chromosome segregation. CRL4 is recruited to chromatin by the replication origin binding protein RepID/DCAF14/PHIP. During mitosis, CRL4 dissociates from RepID and replaces it with RB Binding Protein 7 (RBBP7), which ubiquitinates the SAC mediator BUB3 to enable mitotic exit. During interphase, BUB3 is protected from CRL4-mediated degradation by associating with promyelocytic leukemia (PML) nuclear bodies, ensuring its availability upon mitotic onset. Deficiencies in RepID, CRL4 or RBBP7 delay mitotic exit, increase genomic instability and enhance sensitivity to paclitaxel, a microtubule stabilizer and anti-tumor drug. The spindle assembly checkpoint (SAC) safeguards chromosome segregation by regulating the anaphase promoting complex/cyclosome (APC/C), allowing chromosomes to correctly attach to mitotic spindles. Here the authors reveal a role for Cullin–RING ubiquitin ligase complex 4 (CRL4) in regulating metaphase to anaphase transition via BUB3 degradation.
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Affiliation(s)
- Sang-Min Jang
- Developmental Therapeutics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD, 20892-4255, USA
| | - Jenny F Nathans
- Laboratory of Cancer Biology and Genetics, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD, 20892-4255, USA
| | - Haiqing Fu
- Developmental Therapeutics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD, 20892-4255, USA
| | - Christophe E Redon
- Developmental Therapeutics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD, 20892-4255, USA
| | - Lisa M Jenkins
- Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD, 20892-4255, USA
| | - Bhushan L Thakur
- Developmental Therapeutics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD, 20892-4255, USA
| | - Lőrinc S Pongor
- Developmental Therapeutics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD, 20892-4255, USA
| | - Adrian M Baris
- Developmental Therapeutics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD, 20892-4255, USA
| | - Jacob M Gross
- Developmental Therapeutics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD, 20892-4255, USA
| | - Maura J OʹNeill
- Protein Characterization Laboratory, Cancer Research Technology Program, Frederick National Laboratory for Cancer Research, Frederick, MD, 21701, USA
| | - Fred E Indig
- Confocal Imaging Facility, National Institute on Aging, NIH, Baltimore, MD, 21224, USA
| | - Steven D Cappell
- Laboratory of Cancer Biology and Genetics, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD, 20892-4255, USA
| | - Mirit I Aladjem
- Developmental Therapeutics Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD, 20892-4255, USA.
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CRL4 Ubiquitin Pathway and DNA Damage Response. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2020; 1217:225-239. [PMID: 31898231 DOI: 10.1007/978-981-15-1025-0_14] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/20/2023]
Abstract
DNA damage occurs in a human cell at an average frequency of 10,000 incidences per day by means of external and internal culprits, damage that triggers sequential cellular responses and stalls the cell cycle while activating specific DNA repair pathways. Failure to remove DNA lesions would compromise genomic integrity, leading to human diseases such as cancer and premature aging. If DNA damage is extensive and cannot be repaired, cells undergo apoptosis. DNA damage response (DDR) often entails posttranslational modifications of key DNA repair and DNA damage checkpoint proteins, including phosphorylation and ubiquitination. Cullin-RING ligase 4 (CRL4) enzyme has been found to target multiple DDR proteins for ubiquitination. In this chapter, we will discuss key repair and checkpoint proteins that are subject to ubiquitin-dependent regulation by members of the CRL4 family during ultraviolet light (UV)-induced DNA damage.
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Shi W, Ding R, Zhou PP, Fang Y, Wan R, Chen Y, Jin J. Coordinated Actions Between p97 and Cullin-RING Ubiquitin Ligases for Protein Degradation. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2020; 1217:61-78. [PMID: 31898222 DOI: 10.1007/978-981-15-1025-0_5] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
The cullin-RING ubiquitin ligases comprise the largest subfamily of ubiquitin ligases. They control ubiquitylation and degradation of a large number of protein substrates in eukaryotes. p97 is an ATPase domain-containing protein segregase. It plays essential roles in post-ubiquitylational events in the ubiquitin-proteasome pathway. Together with its cofactors, p97 collaborates with ubiquitin ligases to extract ubiquitylated substrates and deliver them to the proteasome for proteolysis. Here we review the structure, functions, and mechanisms of p97 in cellular protein degradation in coordination with its cofactors and the cullin-RING ubiquitin ligases.
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Affiliation(s)
- Wenbo Shi
- Life Science Institute, Zhejiang University, HangZhou, China
| | - Ran Ding
- Life Science Institute, Zhejiang University, HangZhou, China
| | - Pei Pei Zhou
- Life Science Institute, Zhejiang University, HangZhou, China
| | - Yuan Fang
- Life Science Institute, Zhejiang University, HangZhou, China
| | - Ruixi Wan
- Life Science Institute, Zhejiang University, HangZhou, China
| | - Yilin Chen
- Life Science Institute, Zhejiang University, HangZhou, China
| | - Jianping Jin
- Life Science Institute, Zhejiang University, HangZhou, China.
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Spatial transcriptome profiling by MERFISH reveals subcellular RNA compartmentalization and cell cycle-dependent gene expression. Proc Natl Acad Sci U S A 2019; 116:19490-19499. [PMID: 31501331 PMCID: PMC6765259 DOI: 10.1073/pnas.1912459116] [Citation(s) in RCA: 448] [Impact Index Per Article: 74.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/15/2023] Open
Abstract
The spatial organization of RNAs within cells and spatial patterning of cells within tissues play crucial roles in many biological processes. Here, we demonstrate that multiplexed error-robust FISH (MERFISH) can achieve near-genome-wide, spatially resolved RNA profiling of individual cells with high accuracy and high detection efficiency. Using this approach, we identified RNA species enriched in different subcellular compartments, observed transcriptionally distinct cell states corresponding to different cell-cycle phases, and revealed spatial patterning of transcriptionally distinct cells. Spatially resolved transcriptome quantification within cells further enabled RNA velocity and pseudotime analysis, which revealed numerous genes with cell cycle-dependent expression. We anticipate that spatially resolved transcriptome analysis will advance our understanding of the interplay between gene regulation and spatial context in biological systems. The expression profiles and spatial distributions of RNAs regulate many cellular functions. Image-based transcriptomic approaches provide powerful means to measure both expression and spatial information of RNAs in individual cells within their native environment. Among these approaches, multiplexed error-robust fluorescence in situ hybridization (MERFISH) has achieved spatially resolved RNA quantification at transcriptome scale by massively multiplexing single-molecule FISH measurements. Here, we increased the gene throughput of MERFISH and demonstrated simultaneous measurements of RNA transcripts from ∼10,000 genes in individual cells with ∼80% detection efficiency and ∼4% misidentification rate. We combined MERFISH with cellular structure imaging to determine subcellular compartmentalization of RNAs. We validated this approach by showing enrichment of secretome transcripts at the endoplasmic reticulum, and further revealed enrichment of long noncoding RNAs, RNAs with retained introns, and a subgroup of protein-coding mRNAs in the cell nucleus. Leveraging spatially resolved RNA profiling, we developed an approach to determine RNA velocity in situ using the balance of nuclear versus cytoplasmic RNA counts. We applied this approach to infer pseudotime ordering of cells and identified cells at different cell-cycle states, revealing ∼1,600 genes with putative cell cycle-dependent expression and a gradual transcription profile change as cells progress through cell-cycle stages. Our analysis further revealed cell cycle-dependent and cell cycle-independent spatial heterogeneity of transcriptionally distinct cells. We envision that the ability to perform spatially resolved, genome-wide RNA profiling with high detection efficiency and accuracy by MERFISH could help address a wide array of questions ranging from the regulation of gene expression in cells to the development of cell fate and organization in tissues.
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Cui H, Wang Q, Lei Z, Feng M, Zhao Z, Wang Y, Wei G. DTL promotes cancer progression by PDCD4 ubiquitin-dependent degradation. JOURNAL OF EXPERIMENTAL & CLINICAL CANCER RESEARCH : CR 2019; 38:350. [PMID: 31409387 PMCID: PMC6693180 DOI: 10.1186/s13046-019-1358-x] [Citation(s) in RCA: 77] [Impact Index Per Article: 12.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 05/04/2019] [Accepted: 08/06/2019] [Indexed: 12/20/2022]
Abstract
Background Ubiquitin E3 ligase CUL4A plays important oncogenic roles in the development of cancers. DTL, one of the CUL4-DDB1 associated factors (DCAFs), may involve in the process of cancer development. Programmed cell death 4 (PDCD4) is a tumor suppressor gene involved in cell apoptosis, transformation, invasion and tumor progression. Methods Affinity-purification mass spectrometry was used to identify potential DTL interaction proteins. Co-immunoprecipitation (Co-IP) was performed to verify protein interaction between DTL and PDCD4. mRNA levels in cancer cells and tissues were detected by Quantitative real-time PCR. Lentivirus was used to establish stable overexpression and knocking down cell lines for DTL and PDCD4. Transwell and wound healing assays were used to determine migration ability of cancer cells. Matrigel assay was used to determine invasion ability of cancer cells. MTT and colony formation assays were used to evaluate proliferation of cancer cells. Results In this study, programmed cell death 4 (PDCD4) was identified as a potential substrate of DTL. Co-IP and immunofluorescence assays further confirmed the interaction between DTL and PDCD4. Moreover, DTL overexpression decreased the protein level and accelerated the degradation rate of PDCD4. Through in vitro ubiquitination experiment, we proved that PDCD4 was degraded by DTL through ubiquitination. Clinically DTL was significantly up-regulated in cancer tissues than that in normal tissues. The survival curves showed that cancer patients with higher DTL expression owned lower survival rate. Functional experiments showed that DTL not only enhanced the proliferation and migration abilities of cancer cells, but also promoted the tumorigenesis in nude mice. Rescued experiment results demonstrated that silencing PDCD4 simultaneous with DTL recovered the phenotypes defect caused by DTL knocking down. Conclusions Our results elucidated that DTL enhanced the motility and proliferation of cancer cells through degrading PDCD4 to promote the development of cancers. Electronic supplementary material The online version of this article (10.1186/s13046-019-1358-x) contains supplementary material, which is available to authorized users.
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Affiliation(s)
- Haoran Cui
- Department of Cell Biology and Key Laboratory of Experimental Teratology, Ministry of Education, Shandong University School of Medicine, Jinan, Shandong, China
| | - Qin Wang
- Department of Anesthesiology, Qilu Hospital, Shandong University, Jinan, Shandong, China
| | - Zhenchuan Lei
- College of Marine Life Sciences, Ocean University of China, Qingdao, Shandong, China
| | - Maoxiao Feng
- Department of Cell Biology and Key Laboratory of Experimental Teratology, Ministry of Education, Shandong University School of Medicine, Jinan, Shandong, China
| | - Zhongxi Zhao
- School of Pharmaceutical Sciences, Shandong University, No. 44 West Wenhua Road, Jinan, Shandong, China
| | - Yunshan Wang
- Department of Clinical Laboratory, The Second Hospital of Shandong University, Jinan, Shandong, China.
| | - Guangwei Wei
- Department of Cell Biology and Key Laboratory of Experimental Teratology, Ministry of Education, Shandong University School of Medicine, Jinan, Shandong, China.
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Mazian MA, Suenaga N, Ishii T, Hayashi A, Shiomi Y, Nishitani H. A DNA-binding domain in the C-terminal region of Cdt2 enhances the DNA synthesis-coupled CRL4Cdt2 ubiquitin ligase activity for Cdt1. J Biochem 2019; 165:505-516. [PMID: 30649446 DOI: 10.1093/jb/mvz001] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/25/2018] [Accepted: 01/07/2019] [Indexed: 12/14/2022] Open
Abstract
The Cullin-RING ubiquitin ligase CRL4Cdt2 maintains genome integrity by mediating the cell cycle- and DNA damage-dependent degradation of proteins such as Cdt1, p21 and Set8. Human Cdt2 has two regions, a conserved N-terminal seven WD40 repeat region and a less conserved C-terminal region. Here, we showed that the N-terminal region is sufficient for complex formation with CRL4, but the C-terminal region is required for the full ubiquitin ligase activity. UV irradiation-induced polyubiquitination and degradation of Cdt1 were impaired in Cdt2 (N-terminus only)-expressing cells. Deletion and mutation analysis identified a domain in the C-terminal region that increased ubiquitination activity and displayed DNA-binding activity. The identified domain mediated binding to double-stranded DNA and showed higher affinity binding to single-stranded DNA. As the ligase activity of CRL4Cdt2 depends on proliferating cell nuclear antigen (PCNA) loading onto DNA, the present results suggest that the DNA-binding domain facilitates the CRL4Cdt2-mediated recognition and ubiquitination of substrates bound to PCNA on chromatin.
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Affiliation(s)
- Muadz Ahmad Mazian
- Graduate School of Life Science, University of Hyogo, Kamigori, Akogun Hyogo, Japan
| | - Naohiro Suenaga
- Graduate School of Life Science, University of Hyogo, Kamigori, Akogun Hyogo, Japan
| | - Takashi Ishii
- Graduate School of Life Science, University of Hyogo, Kamigori, Akogun Hyogo, Japan
| | - Akiyo Hayashi
- Graduate School of Life Science, University of Hyogo, Kamigori, Akogun Hyogo, Japan
| | - Yasushi Shiomi
- Graduate School of Life Science, University of Hyogo, Kamigori, Akogun Hyogo, Japan
| | - Hideo Nishitani
- Graduate School of Life Science, University of Hyogo, Kamigori, Akogun Hyogo, Japan
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Yang L, Dai J, Ma M, Mao L, Si L, Cui C, Sheng X, Chi Z, Yu S, Xu T, Yu J, Kong Y, Guo J. Identification of a functional polymorphism within the 3'-untranslated region of denticleless E3 ubiquitin protein ligase homolog associated with survival in acral melanoma. Eur J Cancer 2019; 118:70-81. [PMID: 31325875 DOI: 10.1016/j.ejca.2019.06.006] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/28/2018] [Revised: 05/07/2019] [Accepted: 06/20/2019] [Indexed: 12/25/2022]
Abstract
BACKGROUND High expression of denticleless E3 ubiquitin protein ligase homologue (DTL) correlates with poor disease-free survival and overall survival in cutaneous melanoma, but the molecular features and clinical significance of this gene in acral melanoma (AM) remain unclear. METHODS The expression levels of DTL were compared between AM and benign melanocytic nevi using existing Gene Expression Omnibus data and validated in fresh frozen tissues. Two candidate tag single-nucleotide polymorphisms (SNPs) in the 3'-untranslated region (3'UTR) of DTL in patients with AM were sequenced and analysed for their association with survival in a discovery cohort (n = 570), and the significant SNP was subjected to a replication cohort (n = 201). The expression of DTL was evaluated by immunohistochemistry. The microRNA interacting with rs11275300:C > G was predicted using in silico target prediction tools and validated by in vitro analysis. RESULTS DTL was overexpressed in AM compared with benign melanocytic nevi. rs11275300:C > G was found to be significantly associated with progression-free survival and overall survival of patients with AM in both cohorts and the combined cohort. Furthermore, the DTL expression level in the patients with the rs11275300:G allele was higher than that in patients with the CC genotype. In vitro analysis demonstrated that DTL was a direct target of hsa-miR-4672, and the rs11275300:G allele interfered with the binding affinity of hsa-miR-4672 with the 3'UTR of DTL and thereby increased DTL expression. CONCLUSION The rs11275300:G allele in the 3'UTR of DTL may lead to a poor prognosis and allele-specific increase in the expression of DTL by post-transcriptional regulation in AM.
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Affiliation(s)
- Lu Yang
- Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Renal Cancer and Melanoma, Peking University Cancer Hospital and Institute, Beijing, 100142, China
| | - Jie Dai
- Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Renal Cancer and Melanoma, Peking University Cancer Hospital and Institute, Beijing, 100142, China
| | - Meng Ma
- Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Renal Cancer and Melanoma, Peking University Cancer Hospital and Institute, Beijing, 100142, China; Department of Radiotherapy, Beijing Chest Hospital, Capital Medical University, Beijing Tuberculosis and Thoracic Tumor Research Institute, Beijing, 101149, China
| | - Lili Mao
- Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Renal Cancer and Melanoma, Peking University Cancer Hospital and Institute, Beijing, 100142, China
| | - Lu Si
- Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Renal Cancer and Melanoma, Peking University Cancer Hospital and Institute, Beijing, 100142, China
| | - Chuanliang Cui
- Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Renal Cancer and Melanoma, Peking University Cancer Hospital and Institute, Beijing, 100142, China
| | - Xinan Sheng
- Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Renal Cancer and Melanoma, Peking University Cancer Hospital and Institute, Beijing, 100142, China
| | - Zhihong Chi
- Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Renal Cancer and Melanoma, Peking University Cancer Hospital and Institute, Beijing, 100142, China
| | - Sifan Yu
- Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Renal Cancer and Melanoma, Peking University Cancer Hospital and Institute, Beijing, 100142, China
| | - Tianxiao Xu
- Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Renal Cancer and Melanoma, Peking University Cancer Hospital and Institute, Beijing, 100142, China
| | - Jinyu Yu
- Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Renal Cancer and Melanoma, Peking University Cancer Hospital and Institute, Beijing, 100142, China
| | - Yan Kong
- Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Renal Cancer and Melanoma, Peking University Cancer Hospital and Institute, Beijing, 100142, China.
| | - Jun Guo
- Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Renal Cancer and Melanoma, Peking University Cancer Hospital and Institute, Beijing, 100142, China.
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Prata DP, Costa-Neves B, Cosme G, Vassos E. Unravelling the genetic basis of schizophrenia and bipolar disorder with GWAS: A systematic review. J Psychiatr Res 2019; 114:178-207. [PMID: 31096178 DOI: 10.1016/j.jpsychires.2019.04.007] [Citation(s) in RCA: 60] [Impact Index Per Article: 10.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/20/2018] [Revised: 04/08/2019] [Accepted: 04/10/2019] [Indexed: 01/02/2023]
Abstract
OBJECTIVES To systematically review findings of GWAS in schizophrenia (SZ) and in bipolar disorder (BD); and to interpret findings, with a focus on identifying independent replications. METHOD PubMed search, selection and review of all independent GWAS in SZ or BD, published since March 2011, i.e. studies using non-overlapping samples within each article, between articles, and with those of the previous review (Li et al., 2012). RESULTS From the 22 GWAS included in this review, the genetic associations surviving standard GWAS-significance were for genetic markers in the regions of ACSL3/KCNE4, ADCY2, AMBRA1, ANK3, BRP44, DTL, FBLN1, HHAT, INTS7, LOC392301, LOC645434/NMBR, LOC729457, LRRFIP1, LSM1, MDM1, MHC, MIR2113/POU3F2, NDST3, NKAPL, ODZ4, PGBD1, RENBP, TRANK1, TSPAN18, TWIST2, UGT1A1/HJURP, WHSC1L1/FGFR1 and ZKSCAN4. All genes implicated across both reviews are discussed in terms of their function and implication in neuropsychiatry. CONCLUSION Taking all GWAS to date into account, AMBRA1, ANK3, ARNTL, CDH13, EFHD1 (albeit with different alleles), MHC, PLXNA2 and UGT1A1 have been implicated in either disorder in at least two reportedly non-overlapping samples. Additionally, evidence for a SZ/BD common genetic basis is most strongly supported by the implication of ANK3, NDST3, and PLXNA2.
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Affiliation(s)
- Diana P Prata
- Instituto de Biofísica e Engenharia Biomédica, Faculdade de Ciências, Universidade de Lisboa, Portugal; Centre for Neuroimaging Sciences, Institute of Psychiatry, Psychology & Neuroscience, King's College London, 16 De Crespigny Park, SE5 8AF, UK; Instituto Universitário de Lisboa (ISCTE-IUL), Centro de Investigação e Intervenção Social, Lisboa, Portugal.
| | - Bernardo Costa-Neves
- Lisbon Medical School, University of Lisbon, Av. Professor Egas Moniz, 1649-028, Lisbon, Portugal; Centro Hospitalar Psiquiátrico de Lisboa, Av. do Brasil, 53 1749-002, Lisbon, Portugal
| | - Gonçalo Cosme
- Instituto de Biofísica e Engenharia Biomédica, Faculdade de Ciências, Universidade de Lisboa, Portugal
| | - Evangelos Vassos
- Social, Genetic and Developmental Psychiatry Centre, Institute of Psychiatry, King's College London, 16 De Crespigny Park, SE5 8AF, UK
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Zhang X, Crowley VM, Wucherpfennig TG, Dix MM, Cravatt BF. Electrophilic PROTACs that degrade nuclear proteins by engaging DCAF16. Nat Chem Biol 2019; 15:737-746. [PMID: 31209349 PMCID: PMC6592777 DOI: 10.1038/s41589-019-0279-5] [Citation(s) in RCA: 311] [Impact Index Per Article: 51.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2018] [Accepted: 03/25/2019] [Indexed: 12/18/2022]
Abstract
Ligand-dependent protein degradation has emerged as a compelling strategy to pharmacologically control the protein content of cells. So far, however, only a limited number of E3 ligases have been found to support this process. Here, we use a chemical proteomic strategy that leverages broadly reactive, cysteine-directed electrophilic fragments coupled to selective ligands for intracellular proteins (for example, SLF for FKBP12, JQ1 for BRD4) to screen for heterobifunctional degrader compounds (or proteolysis targeting chimeras, PROTACs) that operate by covalent adduction of E3 ligases. This approach identified DCAF16-a poorly characterized substrate recognition component of CUL4-DDB1 E3 ubiquitin ligases-as a target of electrophilic PROTACs that promote the nuclear-restricted degradation of proteins. We find that only a modest fraction (~10-40%) of DCAF16 needs to be modified to support protein degradation, pointing to the potential for electrophilic PROTACs to induce neosubstrate degradation without substantially perturbing the function of the participating E3 ligase.
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Affiliation(s)
- Xiaoyu Zhang
- The Department of Chemistry and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, CA, USA.
| | - Vincent M Crowley
- The Department of Chemistry and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, CA, USA
| | - Thomas G Wucherpfennig
- The Department of Chemistry and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, CA, USA
| | - Melissa M Dix
- The Department of Chemistry and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, CA, USA
| | - Benjamin F Cravatt
- The Department of Chemistry and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, CA, USA.
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Huang F, Wang M, Liu R, Wang JZ, Schadt E, Haroutunian V, Katsel P, Zhang B, Wang X. CDT2-controlled cell cycle reentry regulates the pathogenesis of Alzheimer's disease. Alzheimers Dement 2019; 15:217-231. [PMID: 30321504 PMCID: PMC6758558 DOI: 10.1016/j.jalz.2018.08.013] [Citation(s) in RCA: 25] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/21/2018] [Revised: 07/07/2018] [Accepted: 08/31/2018] [Indexed: 12/29/2022]
Abstract
INTRODUCTION Altered cell cycle reentry has been observed in Alzheimer's disease (AD). Denticleless (DTL) was predicted as the top driver of a cell cycle subnetwork associated with AD. METHODS We systematically investigated DTL expression in AD and studied the molecular, cellular, and behavioral endophenotypes triggered by DTL overexpression. RESULTS We experimentally validated that CDT2, the protein encoded by DTL, activated cyclin-dependent kinases through downregulating P21, which induced tau hyperphosphorylation and Aβ toxicity, two hallmarks of AD. We demonstrated that cyclin-dependent kinases inhibition by roscovitine not only rescued CDT2-induced cognitive defects but also reversed expression changes induced by DTL overexpression. RNA-seq data from the DTL overexpression experiments revealed the molecular mechanisms underlying CDT2 controlled cell cycle reentry in AD. DISCUSSION These findings provide new insights into the molecular mechanisms of AD pathogenesis and thus pave a way for developing novel therapeutics for AD by targeting AD specific cell cycle networks and drivers.
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Affiliation(s)
- Fang Huang
- Department of Pathophysiology, School of Basic Medicine, Key Laboratory of Education Ministry of China for Neurological Disorders, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Minghui Wang
- Department of Genetics and Genomic Sciences, Mount Sinai Center for Transformative Disease Modeling, Icahn Institute of Genomics and Multiscale Biology, Icahn School of Medicine at Mount Sinai, NY, USA
| | - Rong Liu
- Department of Pathophysiology, School of Basic Medicine, Key Laboratory of Education Ministry of China for Neurological Disorders, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Jian-Zhi Wang
- Department of Pathophysiology, School of Basic Medicine, Key Laboratory of Education Ministry of China for Neurological Disorders, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Eric Schadt
- Department of Genetics and Genomic Sciences, Mount Sinai Center for Transformative Disease Modeling, Icahn Institute of Genomics and Multiscale Biology, Icahn School of Medicine at Mount Sinai, NY, USA
| | - Vahram Haroutunian
- Departments of Psychiatry and Neuroscience, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Department of Psychiatry, JJ Peters VA Medical Center, Bronx, NY, USA; Fishberg Department of Neuroscience, Icahn School of Medicine at Mount Sinai, New York, NY, USA; The Alzheimer's Disease Research Center, Icahn School of Medicine at Mount Sinai, New York NY, USA
| | - Pavel Katsel
- Departments of Psychiatry and Neuroscience, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Bin Zhang
- Department of Genetics and Genomic Sciences, Mount Sinai Center for Transformative Disease Modeling, Icahn Institute of Genomics and Multiscale Biology, Icahn School of Medicine at Mount Sinai, NY, USA.
| | - Xiaochuan Wang
- Department of Pathophysiology, School of Basic Medicine, Key Laboratory of Education Ministry of China for Neurological Disorders, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China; Co-innovation Center of Neuroregeneration, Nantong University, Nantong, JS, China.
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40
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Hayashi A, Giakoumakis NN, Heidebrecht T, Ishii T, Panagopoulos A, Caillat C, Takahara M, Hibbert RG, Suenaga N, Stadnik-Spiewak M, Takahashi T, Shiomi Y, Taraviras S, von Castelmur E, Lygerou Z, Perrakis A, Nishitani H. Direct binding of Cdt2 to PCNA is important for targeting the CRL4 Cdt2 E3 ligase activity to Cdt1. Life Sci Alliance 2018; 1:e201800238. [PMID: 30623174 PMCID: PMC6312923 DOI: 10.26508/lsa.201800238] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/11/2018] [Revised: 12/17/2018] [Accepted: 12/17/2018] [Indexed: 01/18/2023] Open
Abstract
The C-terminal end of Cdt2 contains a PIP box for binding to PCNA to promote CRL4Cdt2 function, creating a new paradigm where the substrate receptor and substrates bind to a common multivalent docking platform for ubiquitination. The CRL4Cdt2 ubiquitin ligase complex is an essential regulator of cell-cycle progression and genome stability, ubiquitinating substrates such as p21, Set8, and Cdt1, via a display of substrate degrons on proliferating cell nuclear antigens (PCNAs). Here, we examine the hierarchy of the ligase and substrate recruitment kinetics onto PCNA at sites of DNA replication. We demonstrate that the C-terminal end of Cdt2 bears a PCNA interaction protein motif (PIP box, Cdt2PIP), which is necessary and sufficient for the binding of Cdt2 to PCNA. Cdt2PIP binds PCNA directly with high affinity, two orders of magnitude tighter than the PIP box of Cdt1. X-ray crystallographic structures of PCNA bound to Cdt2PIP and Cdt1PIP show that the peptides occupy all three binding sites of the trimeric PCNA ring. Mutating Cdt2PIP weakens the interaction with PCNA, rendering CRL4Cdt2 less effective in Cdt1 ubiquitination and leading to defects in Cdt1 degradation. The molecular mechanism we present suggests a new paradigm for bringing substrates to the CRL4-type ligase, where the substrate receptor and substrates bind to a common multivalent docking platform to enable subsequent ubiquitination.
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Affiliation(s)
- Akiyo Hayashi
- Graduate School of Life Science, University of Hyogo, Kamigori, Japan
| | | | - Tatjana Heidebrecht
- Department of Biochemistry, Netherlands Cancer Institute, Amsterdam, The Netherlands
| | - Takashi Ishii
- Graduate School of Life Science, University of Hyogo, Kamigori, Japan
| | | | - Christophe Caillat
- Department of Biochemistry, Netherlands Cancer Institute, Amsterdam, The Netherlands
| | - Michiyo Takahara
- Graduate School of Life Science, University of Hyogo, Kamigori, Japan
| | - Richard G Hibbert
- Department of Biochemistry, Netherlands Cancer Institute, Amsterdam, The Netherlands
| | - Naohiro Suenaga
- Graduate School of Life Science, University of Hyogo, Kamigori, Japan
| | - Magda Stadnik-Spiewak
- Department of Biochemistry, Netherlands Cancer Institute, Amsterdam, The Netherlands
| | | | - Yasushi Shiomi
- Graduate School of Life Science, University of Hyogo, Kamigori, Japan
| | - Stavros Taraviras
- Department of Physiology, School of Medicine, University of Patras, Patras, Greece
| | | | - Zoi Lygerou
- Department of Biology, School of Medicine, University of Patras, Patras, Greece
| | - Anastassis Perrakis
- Department of Biochemistry, Netherlands Cancer Institute, Amsterdam, The Netherlands
| | - Hideo Nishitani
- Graduate School of Life Science, University of Hyogo, Kamigori, Japan
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Fan K, Wang F, Li Y, Chen L, Gao Z, Zhang Y, Duan JY, Huang T, Zhong J, Liu RB, Mao X, Fan H, Guo X, Jin J. CRL4 DCAF2 is required for mature T-cell expansion via Aurora B-regulated proteasome activity. J Autoimmun 2018; 96:74-85. [PMID: 30245026 DOI: 10.1016/j.jaut.2018.08.006] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/05/2018] [Revised: 08/18/2018] [Accepted: 08/21/2018] [Indexed: 12/14/2022]
Abstract
The proliferation of T cells in peripheral lymphoid tissues requires T cell receptor (TCR)-mediated cell cycle entry. However, the underlying mechanism regulating cell cycle progression in mature T cells is incompletely understood. Here, we have identified an E3 ubiquitin ligase, CRL4DCAF2, as a critical mediator controlling M phase exit in activated T cells. DCAF2 expression is induced upon TCR stimulation and its deficiency attenuates T cell expansion. Additionally, DCAF2 T cell-specific knockout mice display impaired peripheral T cell maintenance and reduced severity of various autoimmune diseases. Continuous H4K20me1 modification caused by DCAF2 deficiency inhibits the induction of Aurkb expression, which regulates 26S proteasome activity during G2/M phase. CRL4DCAF2 deficiency causes M phase arrest through proteasome-dependent mechanisms in peripheral T cells. Our findings establish DCAF2 as a novel target for T cell-mediated autoimmunity or inflammatory diseases.
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Affiliation(s)
- Keqi Fan
- Life Sciences Institute, Zhejiang University, Hangzhou, 310058, China
| | - Fei Wang
- Life Sciences Institute, Zhejiang University, Hangzhou, 310058, China
| | - Yiyuan Li
- Life Sciences Institute, Zhejiang University, Hangzhou, 310058, China
| | - Lu Chen
- Life Sciences Institute, Zhejiang University, Hangzhou, 310058, China
| | - Zhengjun Gao
- Life Sciences Institute, Zhejiang University, Hangzhou, 310058, China
| | - Yu Zhang
- Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
| | - Jin-Yuan Duan
- Life Sciences Institute, Zhejiang University, Hangzhou, 310058, China
| | - Tao Huang
- Life Sciences Institute, Zhejiang University, Hangzhou, 310058, China
| | - Jiangyan Zhong
- Life Sciences Institute, Zhejiang University, Hangzhou, 310058, China
| | - Rong-Bei Liu
- Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, 310016, China
| | - Xintao Mao
- Life Sciences Institute, Zhejiang University, Hangzhou, 310058, China
| | - Hengyu Fan
- Life Sciences Institute, Zhejiang University, Hangzhou, 310058, China
| | - Xing Guo
- Life Sciences Institute, Zhejiang University, Hangzhou, 310058, China.
| | - Jin Jin
- Life Sciences Institute, Zhejiang University, Hangzhou, 310058, China; Key Laboratory of Animal Virology of Ministry of Agriculture, Zhejiang University, Hangzhou, 310058, PR China.
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42
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Huang T, Gao Z, Zhang Y, Fan K, Wang F, Li Y, Zhong J, Fan HY, Cao Q, Zhou J, Xiao Y, Hu H, Jin J. CRL4 DCAF2 negatively regulates IL-23 production in dendritic cells and limits the development of psoriasis. J Exp Med 2018; 215:1999-2017. [PMID: 30018073 PMCID: PMC6080916 DOI: 10.1084/jem.20180210] [Citation(s) in RCA: 38] [Impact Index Per Article: 5.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/31/2018] [Revised: 04/18/2018] [Accepted: 06/14/2018] [Indexed: 02/05/2023] Open
Abstract
The E3 ligase CRL4DCAF2 is believed to be a pivotal regulator of the cell cycle and is required for mitotic and S phase progression. The NEDD8-targeting drug MLN4924, which inactivates cullin ring-finger ubiquitin ligases (CRLs), has been examined in clinical trials for various types of lymphoma and acute myeloid leukemia. However, the essential role of CRL4DCAF2 in primary myeloid cells remains poorly understood. MLN4924 treatment, which mimics DCAF2 depletion, also promotes the severity of mouse psoriasis models, consistent with the effects of reduced DCAF2 expression in various autoimmune diseases. Using transcriptomic and immunological approaches, we showed that CRL4DCAF2 in dendritic cells (DCs) regulates the proteolytic fate of NIK and negatively regulates IL-23 production. CRL4DCAF2 promoted the polyubiquitination and subsequent degradation of NIK independent of TRAF3 degradation. DCAF2 deficiency facilitated NIK accumulation and RelB nuclear translocation. DCAF2 DC-conditional knockout mice displayed increased sensitivity to autoimmune diseases. This study shows that CRL4DCAF2 is crucial for controlling NIK stability and highlights a unique mechanism that controls inflammatory diseases.
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Affiliation(s)
- Tao Huang
- Life Sciences Institute, Zhejiang University, Hangzhou, China
| | - Zhengjun Gao
- Life Sciences Institute, Zhejiang University, Hangzhou, China
| | - Yu Zhang
- Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, China
| | - Keqi Fan
- Life Sciences Institute, Zhejiang University, Hangzhou, China
| | - Fei Wang
- Life Sciences Institute, Zhejiang University, Hangzhou, China
| | - Yiyuan Li
- Life Sciences Institute, Zhejiang University, Hangzhou, China
| | - Jiangyan Zhong
- Life Sciences Institute, Zhejiang University, Hangzhou, China
| | - Heng Y Fan
- Life Sciences Institute, Zhejiang University, Hangzhou, China
| | - Qian Cao
- Sir Run Run Shaw Hospital, College of Medicine Zhejiang University, Hangzhou, China
| | - Jiyong Zhou
- Key Laboratory of Animal Virology of Ministry of Agriculture, Zhejiang University, Hangzhou, China
| | - Yichuan Xiao
- Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences/Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Hongbo Hu
- Department of Rheumatology and Immunology, State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, and Collaborative Innovation Center of Biotherapy, Chengdu, China
| | - Jin Jin
- Life Sciences Institute, Zhejiang University, Hangzhou, China
- Key Laboratory of Animal Virology of Ministry of Agriculture, Zhejiang University, Hangzhou, China
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43
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Jang SM, Zhang Y, Utani K, Fu H, Redon CE, Marks AB, Smith OK, Redmond CJ, Baris AM, Tulchinsky DA, Aladjem MI. The replication initiation determinant protein (RepID) modulates replication by recruiting CUL4 to chromatin. Nat Commun 2018; 9:2782. [PMID: 30018425 PMCID: PMC6050238 DOI: 10.1038/s41467-018-05177-6] [Citation(s) in RCA: 28] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/20/2017] [Accepted: 06/07/2018] [Indexed: 12/22/2022] Open
Abstract
Cell cycle progression in mammals is modulated by two ubiquitin ligase complexes, CRL4 and SCF, which facilitate degradation of chromatin substrates involved in the regulation of DNA replication. One member of the CRL4 complex, the WD-40 containing protein RepID (DCAF14/PHIP), selectively binds and activates a group of replication origins. Here we show that RepID recruits the CRL4 complex to chromatin prior to DNA synthesis, thus playing a crucial architectural role in the proper licensing of chromosomes for replication. In the absence of RepID, cells rely on the alternative ubiquitin ligase, SKP2-containing SCF, to progress through the cell cycle. RepID depletion markedly increases cellular sensitivity to SKP2 inhibitors, which triggered massive genome re-replication. Both RepID and SKP2 interact with distinct, non-overlapping groups of replication origins, suggesting that selective interactions of replication origins with specific CRL components execute the DNA replication program and maintain genomic stability by preventing re-initiation of DNA replication. RepID has previously been shown to promote origin firing. Here the authors reveal that RepID regulates replication origins via the recruitment of the CRL4 complex, and prevents re-initiation and unscheduled DNA replication.
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Affiliation(s)
- Sang-Min Jang
- Developmental Therapeutics Branch, Center for Cancer Research, NCI, NIH, Bethesda, MD, 20892-4255, USA
| | - Ya Zhang
- Developmental Therapeutics Branch, Center for Cancer Research, NCI, NIH, Bethesda, MD, 20892-4255, USA
| | - Koichi Utani
- Developmental Therapeutics Branch, Center for Cancer Research, NCI, NIH, Bethesda, MD, 20892-4255, USA
| | - Haiqing Fu
- Developmental Therapeutics Branch, Center for Cancer Research, NCI, NIH, Bethesda, MD, 20892-4255, USA
| | - Christophe E Redon
- Developmental Therapeutics Branch, Center for Cancer Research, NCI, NIH, Bethesda, MD, 20892-4255, USA
| | - Anna B Marks
- Developmental Therapeutics Branch, Center for Cancer Research, NCI, NIH, Bethesda, MD, 20892-4255, USA
| | - Owen K Smith
- Developmental Therapeutics Branch, Center for Cancer Research, NCI, NIH, Bethesda, MD, 20892-4255, USA
| | - Catherine J Redmond
- Developmental Therapeutics Branch, Center for Cancer Research, NCI, NIH, Bethesda, MD, 20892-4255, USA
| | - Adrian M Baris
- Developmental Therapeutics Branch, Center for Cancer Research, NCI, NIH, Bethesda, MD, 20892-4255, USA
| | - Danielle A Tulchinsky
- Developmental Therapeutics Branch, Center for Cancer Research, NCI, NIH, Bethesda, MD, 20892-4255, USA
| | - Mirit I Aladjem
- Developmental Therapeutics Branch, Center for Cancer Research, NCI, NIH, Bethesda, MD, 20892-4255, USA.
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44
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Regulation of Mammalian DNA Replication via the Ubiquitin-Proteasome System. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2018; 1042:421-454. [PMID: 29357069 DOI: 10.1007/978-981-10-6955-0_19] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/09/2023]
Abstract
Proper regulation of DNA replication ensures the faithful transmission of genetic material essential for optimal cellular and organismal physiology. Central to this regulation is the activity of a set of enzymes that induce or reverse posttranslational modifications of various proteins critical for the initiation, progression, and termination of DNA replication. This is particularly important when DNA replication proceeds in cancer cells with elevated rates of genomic instability and increased proliferative capacities. Here, we describe how DNA replication in mammalian cells is regulated via the activity of the ubiquitin-proteasome system as well as the consequence of derailed ubiquitylation signaling involved in this important cellular activity.
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45
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Identification of differentially accumulated proteins involved in regulating independent and combined osmosis and cadmium stress response in Brachypodium seedling roots. Sci Rep 2018; 8:7790. [PMID: 29773844 PMCID: PMC5958118 DOI: 10.1038/s41598-018-25959-8] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/22/2017] [Accepted: 05/01/2018] [Indexed: 12/24/2022] Open
Abstract
In this study, we aimed to identify differentially accumulated proteins (DAPs) involved in PEG mock osmotic stress, cadmium (Cd2+) stress, and their combined stress responses in Brachypodium distachyon seedling roots. The results showed that combined PEG and Cd2+ stresses had more significant effects on Brachypodium seedling root growth, physiological traits, and ultrastructures when compared with each individual stress. Totally, 106 DAPs were identified that are responsive to individual and combined stresses in roots. These DAPs were mainly involved in energy metabolism, detoxification and stress defense and protein metabolism. Principal component analysis revealed that DAPs from Cd2+ and combined stress treatments were grouped closer than those from osmotic stress treatment, indicating that Cd2+ and combined stresses had more severe influences on the root proteome than osmotic stress alone. Protein-protein interaction analyses highlighted a 14-3-3 centered sub-network that synergistically responded to osmotic and Cd2+ stresses and their combined stresses. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis of 14 key DAP genes revealed that most genes showed consistency between transcriptional and translational expression patterns. A putative pathway of proteome metabolic changes in Brachypodium seedling roots under different stresses was proposed, which revealed a complicated synergetic responsive network of plant roots to adverse environments.
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46
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Vanderdys V, Allak A, Guessous F, Benamar M, Read PW, Jameson MJ, Abbas T. The Neddylation Inhibitor Pevonedistat (MLN4924) Suppresses and Radiosensitizes Head and Neck Squamous Carcinoma Cells and Tumors. Mol Cancer Ther 2018; 17:368-380. [PMID: 28838998 PMCID: PMC5805645 DOI: 10.1158/1535-7163.mct-17-0083] [Citation(s) in RCA: 50] [Impact Index Per Article: 7.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2017] [Revised: 07/06/2017] [Accepted: 08/17/2017] [Indexed: 11/16/2022]
Abstract
The cullin RING E3 ubiquitin ligase 4 (CRL4) with its substrate receptor CDT2 (CRL4-CDT2) is emerging as a critical regulator of DNA replication through targeting CDT1, SET8, and p21 for ubiquitin-dependent proteolysis. The aberrant increased stability of these proteins in cells with inactivated CRL4-CDT2 results in DNA rereplication, which is deleterious to cells due to the accumulation of replication intermediates and stalled replication forks. Here, we demonstrate that CDT2 is overexpressed in head and neck squamous cell carcinoma (HNSCC), and its depletion by siRNA inhibits the proliferation of human papilloma virus-negative (HPV-ve) HNSCC cells primarily through the induction of rereplication. Treatment of HNSCC with the NEDD8-activating enzyme inhibitor pevonedistat (MLN4924), which inhibits all cullin-based ligases, induces significant rereplication and inhibits HNSCC cell proliferation in culture and HNSCC xenografts in mice. Pevonedistat additionally sensitizes HNSCC cells to ionizing radiation (IR) and enhances IR-induced suppression of xenografts in mice. Induction of rereplication via CDT2 depletion, or via the stabilization or activation of CDT1, also radiosensitizes HNSCC cells. Collectively, these results demonstrate that induction of rereplication represents a novel approach to treating radioresistant HNSCC tumors and suggest that pevonedistat may be considered as an adjuvant for IR-based treatments. Mol Cancer Ther; 17(2); 368-80. ©2017 AACRSee all articles in this MCT Focus section, "Developmental Therapeutics in Radiation Oncology."
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Affiliation(s)
- Vanessa Vanderdys
- Department of Radiation Oncology, University of Virginia, Charlottesville, Virginia
| | - Amir Allak
- Department of Radiation Oncology, University of Virginia, Charlottesville, Virginia
- Department of Otolaryngology, Head and Neck Surgery, University of Virginia, Charlottesville, Virginia
| | - Fadila Guessous
- Department of Radiation Oncology, University of Virginia, Charlottesville, Virginia
| | - Mouadh Benamar
- Department of Radiation Oncology, University of Virginia, Charlottesville, Virginia
- Department of Biochemistry and Molecular Genetics, University of Virginia, Charlottesville, Virginia
| | - Paul W Read
- Department of Radiation Oncology, University of Virginia, Charlottesville, Virginia
| | - Mark J Jameson
- Department of Otolaryngology, Head and Neck Surgery, University of Virginia, Charlottesville, Virginia
| | - Tarek Abbas
- Department of Radiation Oncology, University of Virginia, Charlottesville, Virginia.
- Department of Biochemistry and Molecular Genetics, University of Virginia, Charlottesville, Virginia
- Center for Cell Signaling, University of Virginia, Charlottesville, Virginia
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47
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von Kopylow K, Spiess AN. Human spermatogonial markers. Stem Cell Res 2017; 25:300-309. [PMID: 29239848 DOI: 10.1016/j.scr.2017.11.011] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/01/2016] [Revised: 11/06/2017] [Accepted: 11/13/2017] [Indexed: 12/22/2022] Open
Abstract
In this review, we provide an up-to-date compilation of published human spermatogonial markers, with focus on the three nuclear subtypes Adark, Apale and B. In addition, we have extended our recently published list of putative spermatogonial markers with protein expression and RNA-sequencing data from the Human Protein Atlas and supported these by literature evidence. Most importantly, we have put substantial effort in acquiring a comprehensive list of new and potentially interesting markers by refiltering the raw data of 15 published germ cell expression datasets (four human, eleven rodent) and subsequent building of intersections to acquire a robust, cross-species set of spermatogonia-enriched or -specific transcripts.
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Affiliation(s)
- Kathrein von Kopylow
- Department of Andrology, University Hospital Hamburg-Eppendorf, Hamburg, Germany.
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48
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Yan H, Bi L, Wang Y, Zhang X, Hou Z, Wang Q, Snijders AM, Mao JH. Integrative analysis of multi-omics data reveals distinct impacts of DDB1-CUL4 associated factors in human lung adenocarcinomas. Sci Rep 2017; 7:333. [PMID: 28336923 PMCID: PMC5428704 DOI: 10.1038/s41598-017-00512-1] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2016] [Accepted: 02/28/2017] [Indexed: 12/22/2022] Open
Abstract
Many DDB1-CUL4 associated factors (DCAFs) have been identified and serve as substrate receptors. Although the oncogenic role of CUL4A has been well established, specific DCAFs involved in cancer development remain largely unknown. Here we infer the potential impact of 19 well-defined DCAFs in human lung adenocarcinomas (LuADCs) using integrative omics analyses, and discover that mRNA levels of DTL, DCAF4, 12 and 13 are consistently elevated whereas VBRBP is reduced in LuADCs compared to normal lung tissues. The transcriptional levels of DCAFs are significantly correlated with their gene copy number variations. SKIP2, DTL, DCAF6, 7, 8, 13 and 17 are frequently gained whereas VPRBP, PHIP, DCAF10, 12 and 15 are frequently lost. We find that only transcriptional level of DTL is robustly, significantly and negatively correlated with overall survival across independent datasets. Moreover, DTL-correlated genes are enriched in cell cycle and DNA repair pathways. We also identified that the levels of 25 proteins were significantly associated with DTL overexpression in LuADCs, which include significant decreases in protein level of the tumor supressor genes such as PDCD4, NKX2-1 and PRKAA1. Our results suggest that different CUL4-DCAF axis plays the distinct roles in LuADC development with possible relevance for therapeutic target development.
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Affiliation(s)
- Hong Yan
- Department of Laboratory Medicine, Nanjing Chest Hospital, 215 Guangzhou Road, Nanjing, 210029, China
- Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA
| | - Lei Bi
- Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA
- School of Preclinical Medicine, Nanjing University of Chinese Medicine, 138 Xianlin Road, Nanjing, 210023, China
| | - Yunshan Wang
- Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA
- International Biotechnology R&D Center, Shandong University School of Ocean, Weihai, Shandong, 264209, China
| | - Xia Zhang
- Department of Tuberculosis, Nanjing Chest Hospital, 215 Guangzhou Road, Nanjing, 210029, China
| | - Zhibo Hou
- Department of First Respiratory, Nanjing Chest Hospital, 215 Guangzhou Road, Nanjing, 210029, China
| | - Qian Wang
- Department of Laboratory Medicine, Nanjing Chest Hospital, 215 Guangzhou Road, Nanjing, 210029, China
| | - Antoine M Snijders
- Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA
| | - Jian-Hua Mao
- Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA.
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Overexpression of denticleless E3 ubiquitin protein ligase homolog (DTL) is related to poor outcome in gastric carcinoma. Oncotarget 2017; 6:36615-24. [PMID: 26472028 PMCID: PMC4742199 DOI: 10.18632/oncotarget.5620] [Citation(s) in RCA: 33] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/13/2015] [Accepted: 10/02/2015] [Indexed: 12/17/2022] Open
Abstract
BACKGROUND Denticleless E3 ubiquitin protein ligase homolog (DTL) has been identified in amplified region (1q32) of several cancers and has an oncogenic function. In this study, we tested whether DTL acts as a cancer-promoting gene through its activation/overexpression in gastric cancer (GC). METHODS We analyzed 7 GC cell lines and 100 primary tumors that were curatively resected in our hospital between 2001 and 2003. RESULTS Overexpression of the DTL protein was detected in GC cell lines (4/7 cell lines; 57%) and primary GC tumor samples (42/100 cases; 42%). Knockdown of DTL using several specific siRNAs inhibited the proliferation, migration and invasion in a TP53 mutation-independent manner. Overexpression of the DTL was significantly correlated with lymphatic invasion, deeper tumor depth and higher recurrence rate. Patients with DTL-overexpressing tumors had a worse survival rate than those with non-expressing tumors in overall survival (P = 0.0498, log-rank test) and disease-free survival (P = 0.0324, log-rank test). In a multivariate analysis, DTL positivity was independently associated with a worse overall survival (P = 0.0104, hazard ratio 3.7 [1.36-10.1]) and disease-free survival (P = 0.0070 (hazard ratio, 3.9 (1.45-10.46)) following radical gastrectomy. CONCLUSIONS These findings suggest that DTL overexpression plays a crucial role in tumor cell proliferation and highlights its usefulness as a prognosticator and potential therapeutic target in gastric cancer.
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Tanaka M, Takahara M, Nukina K, Hayashi A, Sakai W, Sugasawa K, Shiomi Y, Nishitani H. Mismatch repair proteins recruited to ultraviolet light-damaged sites lead to degradation of licensing factor Cdt1 in the G1 phase. Cell Cycle 2017; 16:673-684. [PMID: 28278049 DOI: 10.1080/15384101.2017.1295179] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/23/2023] Open
Abstract
Cdt1 is rapidly degraded by CRL4Cdt2 E3 ubiquitin ligase after UV (UV) irradiation. Previous reports revealed that the nucleotide excision repair (NER) pathway is responsible for the rapid Cdt1-proteolysis. Here, we show that mismatch repair (MMR) proteins are also involved in the degradation of Cdt1 after UV irradiation in the G1 phase. First, compared with the rapid (within ∼15 min) degradation of Cdt1 in normal fibroblasts, Cdt1 remained stable for ∼30 min in NER-deficient XP-A cells, but was degraded within ∼60 min. The delayed degradation was also dependent on PCNA and CRL4Cdt2. The MMR proteins Msh2 and Msh6 were recruited to the UV-damaged sites of XP-A cells in the G1 phase. Depletion of these factors with small interfering RNAs prevented Cdt1 degradation in XP-A cells. Similar to the findings in XP-A cells, depletion of XPA delayed Cdt1 degradation in normal fibroblasts and U2OS cells, and co-depletion of Msh6 further prevented Cdt1 degradation. Furthermore, depletion of Msh6 alone delayed Cdt1 degradation in both cell types. When Cdt1 degradation was attenuated by high Cdt1 expression, repair synthesis at the damaged sites was inhibited. Our findings demonstrate that UV irradiation induces multiple repair pathways that activate CRL4Cdt2 to degrade its target proteins in the G1 phase of the cell cycle, leading to efficient repair of DNA damage.
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Affiliation(s)
- Miyuki Tanaka
- a Graduate School of Life Science , University of Hyogo , Kamigori, Ako-gun , Hyogo , Japan
| | - Michiyo Takahara
- a Graduate School of Life Science , University of Hyogo , Kamigori, Ako-gun , Hyogo , Japan
| | - Kohei Nukina
- a Graduate School of Life Science , University of Hyogo , Kamigori, Ako-gun , Hyogo , Japan
| | - Akiyo Hayashi
- a Graduate School of Life Science , University of Hyogo , Kamigori, Ako-gun , Hyogo , Japan
| | - Wataru Sakai
- b Biosignal Research Center , Kobe University , Kobe , Hyogo , Japan
| | - Kaoru Sugasawa
- b Biosignal Research Center , Kobe University , Kobe , Hyogo , Japan
| | - Yasushi Shiomi
- a Graduate School of Life Science , University of Hyogo , Kamigori, Ako-gun , Hyogo , Japan
| | - Hideo Nishitani
- a Graduate School of Life Science , University of Hyogo , Kamigori, Ako-gun , Hyogo , Japan
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