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Kumar A. ZINC1797251, a novel natural product small molecule targets viral oncoprotein E6 in human papillomavirus-16 positive cervical cancer cells. J Biomol Struct Dyn 2025:1-13. [PMID: 40313056 DOI: 10.1080/07391102.2025.2497459] [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: 12/08/2023] [Accepted: 05/03/2024] [Indexed: 05/03/2025]
Abstract
Cervical cancer burden due to recurrent human papillomavirus (HPV) infections necessitates the urgent need to impede viral proliferation targeting the oncogene E6 of the high-risk serotype HPV16. This study aims to identify a small molecule from a natural product library that could prevent a tumorigenic complex of E6 with p53 in HPV16-positive cervical cancer cells. In silico methods such as high-throughput virtual screening (HTVS) of natural product like library ZINC database followed by atomistic molecular dynamics (MD) simulations were performed to identify lead natural compound. This was validated with in vitro analysis using HPV16 positive SiHa cells and CaSki cells by MTT and flow cytometry assays. Virtual screening identified top 10 compounds with high affinity for HPV16 E6. The docking scores, Protein-Ligand Interaction Profiler analysis, MD simulation and molecular mechanics Poisson Boltzmann surface area-based binding energy estimation narrowed down the search to ZINC1797251, a molecule with stable binding, low energy scores and consistent H-bonds, establishing that it could prevent interaction of p53 and E6. ZINC1797251 inhibited the proliferation of SiHa and CaSki cells with a GI50 values of 615.40 and 417.30 nM, respectively. The compound reduced HPV16 E6, while increased p53 positive populations in SiHa and CaSki cells. Treatment with ZINC1797251 induced the G1 cell phase arrest and promoted early and late phase apoptosis in these cells. The restoration of tumor inhibitory activity of p53 in HPV-infected cervical cancer cells to promote apoptosis could be achieved using the ZINC molecule-ZINC1797251. However, further studies are deemed essential for further developments.
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Affiliation(s)
- Ashish Kumar
- Department of Microbiology & Clinical Parasitology, College of Medicine, King Khalid University, Abha, Saudi Arabia
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2
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Mishan MA, Choo YM, Winkler J, Hamann MT, Karan D. Manzamine A: A promising marine-derived cancer therapeutic for multi-targeted interactions with E2F8, SIX1, AR, GSK-3β, and V-ATPase - A systematic review. Eur J Pharmacol 2025; 990:177295. [PMID: 39863145 DOI: 10.1016/j.ejphar.2025.177295] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/12/2024] [Revised: 01/21/2025] [Accepted: 01/22/2025] [Indexed: 01/27/2025]
Abstract
Manzamine A, a natural compound derived from various sponge genera, features a β-carboline structure and exhibits a range of biological activities, including anti-inflammatory and antimalarial effects. Its potential as an anticancer agent has been explored in several tumor models, both in vitro and in vivo, showing effects through mechanisms such as cytotoxicity, regulation of the cell cycle, inhibition of cell migration, epithelial-to-mesenchymal transition (EMT), autophagy, and apoptosis through multi-target interactions of E2F transcriptional factors, ribosomal S6 kinases, androgen receptor (AR), SIX1, GSK-3β, v-ATPase, and p53/p21/p27 cascades. This systematic review evaluates existing literature on the potential application of this marine alkaloid as a novel cancer therapy, highlighting its promising ability to inhibit cancer cell growth while causing minimal side effects.
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Affiliation(s)
- Mohammad Amir Mishan
- Department of Urology, Brown Cancer Center, 505 S Hancock Street, Louisville, KY, USA
| | - Yeun-Mun Choo
- Department of Chemistry, Faculty of Science, University of Malaya, 50603, Kuala Lumpur, Malaysia
| | - Jeffery Winkler
- Department of Chemistry, The University of Pennsylvania, Philadelphia, PA, USA
| | - Mark T Hamann
- Department of Drug Discovery and Biomedical Sciences and Public Health, Colleges of Pharmacy and Medicine, Hollings Cancer Center, Medical University of South Carolina, Charleston, SC, USA
| | - Dev Karan
- Department of Urology, Brown Cancer Center, 505 S Hancock Street, Louisville, KY, USA.
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3
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Avenhaus A, Velimirović M, Bulkescher J, Scheffner M, Hoppe-Seyler F, Hoppe-Seyler K. E6AP is essential for the proliferation of HPV-positive cancer cells by preventing senescence. PLoS Pathog 2025; 21:e1012914. [PMID: 39919145 PMCID: PMC11805377 DOI: 10.1371/journal.ppat.1012914] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/23/2024] [Accepted: 01/16/2025] [Indexed: 02/09/2025] Open
Abstract
Oncogenic types of human papillomaviruses (HPVs) are major human carcinogens. The formation of a trimeric complex between the HPV E6 oncoprotein, the cellular ubiquitin ligase E6AP and the p53 tumor suppressor protein leads to proteolytic p53 degradation and plays a central role for HPV-induced cell transformation. We here uncover that E6AP silencing in HPV-positive cancer cells ultimately leads to efficient induction of cellular senescence, revealing that E6AP acts as a potent anti-senescent factor in these cells. Thus, although the downregulation of either E6 or E6AP expression also acts partially pro-apoptotic, HPV-positive cancer cells surviving E6 repression proliferate further, whereas they become irreversibly growth-arrested upon E6AP repression. We moreover show that the senescence induction following E6AP downregulation is mechanistically highly dependent on induction of the p53/p21 axis, other than the known pro-senescent response of HPV-positive cancer cells following combined downregulation of the viral E6 and E7 oncoproteins. Of further note, repression of E6AP allows senescence induction in the presence of the anti-senescent HPV E7 protein. Yet, despite these mechanistic differences, the pathways underlying the pro-senescent effects of E6AP or E6/E7 repression ultimately converge by being both dependent on the cellular pocket proteins pRb and p130. Taken together, our results uncover a hitherto unrecognized and potent anti-senescent function of the E6AP protein in HPV-positive cancer cells, which is essential for their sustained proliferation. Our results further indicate that interfering with E6AP expression or function could result in therapeutically desired effects in HPV-positive cancer cells by efficiently inducing an irreversible growth arrest. Since the critical role of the E6/E6AP/p53 complex for viral transformation is conserved between different oncogenic HPV types, this approach could provide a therapeutic strategy, which is not HPV type-specific.
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Affiliation(s)
- Alicia Avenhaus
- German Cancer Research Center (DKFZ), Molecular Therapy of Virus-Associated Cancers, Heidelberg, Germany
- Faculty of Biosciences, Heidelberg University, Heidelberg, Germany
| | - Milica Velimirović
- German Cancer Research Center (DKFZ), Molecular Therapy of Virus-Associated Cancers, Heidelberg, Germany
- Faculty of Biosciences, Heidelberg University, Heidelberg, Germany
| | - Julia Bulkescher
- German Cancer Research Center (DKFZ), Molecular Therapy of Virus-Associated Cancers, Heidelberg, Germany
| | - Martin Scheffner
- Department of Biology, Konstanz Research School Chemical Biology, University of Konstanz, Konstanz, Germany
| | - Felix Hoppe-Seyler
- German Cancer Research Center (DKFZ), Molecular Therapy of Virus-Associated Cancers, Heidelberg, Germany
| | - Karin Hoppe-Seyler
- German Cancer Research Center (DKFZ), Molecular Therapy of Virus-Associated Cancers, Heidelberg, Germany
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4
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Tachibana S, Otaki Y, Watanabe T, Goto J, Ochi H, Tanaka T, Ono H, Yamaguchi R, Sato J, Takahashi H, Arimoto T, Goto K, Watanabe M. Diacylglycerol Kinase ζ Attenuates Doxorubicin-Induced Cardiotoxicity Through p53 Degradation. J Am Heart Assoc 2025; 14:e035608. [PMID: 39719406 PMCID: PMC12054516 DOI: 10.1161/jaha.124.035608] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/04/2024] [Accepted: 08/20/2024] [Indexed: 12/26/2024]
Abstract
BACKGROUND Doxorubicin-induced cardiotoxicity is still an important medical problem associated with a high mortality rate in cancer survivors. p53 plays a key role in doxorubicin-induced cardiotoxicity. Diacylglycerol kinase ζ (Dgkζ), a 130-kDa enzyme abundant in cardiomyocytes, regulates the p53 protein expression level in neurons. To elucidate the mechanism of doxorubicin-induced cardiotoxicity, we focused on the functional role of Dgkζ and its interaction with heat shock protein 70 (Hsp70)-related ubiquitin E3 ligases such as E6-associated protein (E6ap) and C-terminus of Hsp70-interacting protein. METHODS AND RESULTS Protein interactions of Dgkζ with Hsp70 and E6ap were confirmed by immunoprecipitation, but not C-terminus of Hsp70-interacting protein. We administered doxorubicin in cardiac-specific overexpression of Dgkζ transgenic (Dgkζ-Tg) mice and wild-type littermates. Dgkζ-Tg mice showed lower p53 protein expression levels, preserved cardiac function, and improved survival rates compared with wild-type littermates after doxorubicin administration. RNA sequence analysis of myocardial tissues from Dgkζ-Tg after doxorubicin stimulation identified Hspa1b encoding Hsp70 as the differentially expressed gene. Dgkζ overexpression increased proteasomal p53 degradation and attenuated cardiomyocyte apoptosis after doxorubicin stimulation in cardiomyocytes, which was reversed by knockdown of E6ap. Dgkζ interacted with E6ap through ankyrin-like repeats. The overexpression of mutant Dgkζ, lacking ankyrin-like repeats, failed to inhibit p53 protein expression after doxorubicin stimulation. In Dgkζ-overexpressing cardiomyocytes, expression levels of p53 and caspase-3 were increased by knockdown of the C-terminus of Hsp70-interacting protein. CONCLUSIONS We demonstrated for the first time that Dgkζ augments p53 ubiquitin-proteasome degradation and ameliorates doxorubicin-induced cardiotoxicity by interacting with Hsp70 and E3 ligases such as E6ap and C-terminus of Hsp70-interacting protein.
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Affiliation(s)
- Shingo Tachibana
- Department of Cardiology, Pulmonology, and NephrologyYamagata University School of MedicineYamagataJapan
| | - Yoichiro Otaki
- Department of Cardiology, Pulmonology, and NephrologyYamagata University School of MedicineYamagataJapan
| | - Tetsu Watanabe
- Department of Cardiology, Pulmonology, and NephrologyYamagata University School of MedicineYamagataJapan
| | - Jun Goto
- Department of Cardiology, Pulmonology, and NephrologyYamagata University School of MedicineYamagataJapan
| | - Haruki Ochi
- Institute for Promotion of Medical Science Research, Faculty of MedicineYamagata UniversityYamagataJapan
| | - Toshiaki Tanaka
- Department of Anatomy and Cell Biology, School of MedicineYamagata UniversityYamagataJapan
| | - Hiroe Ono
- Department of Cardiology, Pulmonology, and NephrologyYamagata University School of MedicineYamagataJapan
| | - Ryuhei Yamaguchi
- Department of Cardiology, Pulmonology, and NephrologyYamagata University School of MedicineYamagataJapan
| | - Junya Sato
- Department of Cardiology, Pulmonology, and NephrologyYamagata University School of MedicineYamagataJapan
| | - Hiroki Takahashi
- Department of Cardiology, Pulmonology, and NephrologyYamagata University School of MedicineYamagataJapan
| | - Takanori Arimoto
- Department of Cardiology, Pulmonology, and NephrologyYamagata University School of MedicineYamagataJapan
| | - Kaoru Goto
- Department of Anatomy and Cell Biology, School of MedicineYamagata UniversityYamagataJapan
| | - Masafumi Watanabe
- Department of Cardiology, Pulmonology, and NephrologyYamagata University School of MedicineYamagataJapan
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Lin Z, Assaraf YG, Kwok HF. Peptides for microbe-induced cancers: latest therapeutic strategies and their advanced technologies. Cancer Metastasis Rev 2024; 43:1315-1336. [PMID: 39008152 DOI: 10.1007/s10555-024-10197-4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/26/2023] [Accepted: 06/14/2024] [Indexed: 07/16/2024]
Abstract
Cancer is a significant global health concern associated with multiple distinct factors, including microbial and viral infections. Numerous studies have elucidated the role of microorganisms, such as Helicobacter pylori (H. pylori), as well as viruses for example human papillomavirus (HPV), hepatitis B virus (HBV), and hepatitis C virus (HCV), in the development of human malignancies. Substantial attention has been focused on the treatment of these microorganism- and virus-associated cancers, with promising outcomes observed in studies employing peptide-based therapies. The current paper provides an overview of microbe- and virus-induced cancers and their underlying molecular mechanisms. We discuss an assortment of peptide-based therapies which are currently being developed, including tumor-targeting peptides and microbial/viral peptide-based vaccines. We describe the major technological advancements that have been made in the design, screening, and delivery of peptides as anticancer agents. The primary focus of the current review is to provide insight into the latest research and development in this field and to provide a realistic glimpse into the future of peptide-based therapies for microbe- and virus-induced neoplasms.
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Affiliation(s)
- Ziqi Lin
- Cancer Centre, Faculty of Health Sciences, University of Macau, Avenida da Universidade, Taipa, Macau SAR
- Department of Biomedical Sciences, Faculty of Health Sciences, University of Macau, Avenida da Universidade, Taipa, Macau SAR
| | - Yehuda G Assaraf
- The Fred Wyszkowski Cancer Research Lab, Faculty of Biology, Technion-Israel Instituteof Technology, Haifa, 3200003, Israel
| | - Hang Fai Kwok
- Cancer Centre, Faculty of Health Sciences, University of Macau, Avenida da Universidade, Taipa, Macau SAR.
- Department of Biomedical Sciences, Faculty of Health Sciences, University of Macau, Avenida da Universidade, Taipa, Macau SAR.
- MoE Frontiers Science Center for Precision Oncology, University of Macau, Avenida de Universidade, Taipa, Macau SAR.
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Kumar A. Human papillomavirus-16 E6-positive cervical cancer attenuated by potent 2-(4-biphenylyl)-N-(1-ethyl-4-piperidinyl) acetamide second-generation analogs with improved binding affinity. Biotechnol Appl Biochem 2024; 71:1428-1439. [PMID: 39039663 DOI: 10.1002/bab.2639] [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: 04/17/2024] [Accepted: 07/06/2024] [Indexed: 07/24/2024]
Abstract
Human papillomavirus (HPV) infection, particularly HPV16, is a major contributor to the development of cervical cancer. Given the urgent need for novel therapeutic strategies targeting HPV-associated cancers, this study focuses on characterizing second-generation analogs of a lead compound, as a potential inhibitor of HPV16-E6. Protein-ligand docking, Gibbs binding free energy estimation, and molecular dynamics simulations were conducted. HPV16-infected SiHa and CaSki cell lines were used. MTT (3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide) assay for proliferation and flow cytometry for target inhibition and apoptosis were employed. Computational and cell proliferation analyses revealed that modifications to E6-855, particularly in the piperidinyl group, enhanced binding affinities against HPV16-E6, with E6-272 demonstrating superior binding properties. Molecular dynamics simulations confirmed the stable binding of E6-272 to HPV16-E6, supported by favorable binding energy estimates. E6-272 inhibited the proliferation of SiHa and CaSki cells with GI50 values of 32.56 and 62.09 nM, respectively. The compound reduced HPV16-E6-positive population, while inducing the early and late phase apoptosis in these cells. Structural alterations at the piperidinyl group of E6-855 identified E6-272 as a promising inhibitor of HPV16-E6 with improved efficacy against HPV16-E6. Further experimental validation of E6-272 and its analogs warrant to advance its clinical utility in combating HPV-associated cancers.
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Affiliation(s)
- Ashish Kumar
- Department of Microbiology & Clinical Parasitology, College of Medicine, King Khalid University, Abha, Saudi Arabia
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7
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Blanco R, Muñoz JP. HPV and HCMV in Cervical Cancer: A Review of Their Co-Occurrence in Premalignant and Malignant Lesions. Viruses 2024; 16:1699. [PMID: 39599814 PMCID: PMC11599080 DOI: 10.3390/v16111699] [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: 09/25/2024] [Revised: 10/24/2024] [Accepted: 10/26/2024] [Indexed: 11/29/2024] Open
Abstract
Cervical cancer remains a significant global health concern, particularly in low- and middle-income countries. While persistent infection with high-risk human papillomavirus (HR-HPV) is essential for cervical cancer development, it is not sufficient on its own, suggesting the involvement of additional cofactors. The human cytomegalovirus (HCMV) is a widespread β-herpesvirus known for its ability to establish lifelong latency and reactivate under certain conditions, often contributing to chronic inflammation and immune modulation. Emerging evidence suggests that HCMV may play a role in various cancers, including cervical cancer, through its potential to influence oncogenic pathways and disrupt host immune responses. This review explores clinical evidence regarding the co-presence of HR-HPV and HCMV in premalignant lesions and cervical cancer. The literature reviewed indicates that HCMV is frequently detected in cervical lesions, particularly in those co-infected with HPV, suggesting a potential synergistic interaction that could enhance HPV's oncogenic effects, thereby facilitating the progression from low-grade squamous intraepithelial lesions (LSIL) to high-grade squamous intraepithelial lesions (HSIL) and invasive cancer. Although the precise molecular mechanisms were not thoroughly investigated in this review, the clinical evidence suggests the importance of considering HCMV alongside HPV in the management of cervical lesions. A better understanding of the interaction between HR-HPV and HCMV may lead to improved diagnostic, therapeutic, and preventive strategies for cervical cancer.
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Affiliation(s)
- Rancés Blanco
- Independent Researcher, Av. Vicuña Mackenna Poniente 6315, La Florida 8240000, Chile
| | - Juan P. Muñoz
- Laboratorio de Bioquímica, Departamento de Química, Facultad de Ciencias, Universidad de Tarapacá, Arica 1000007, Chile
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8
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Zhang A, Zheng X, Chen S, Duan G. In vitro study of HPV18-positive cervical cancer HeLa cells based on CRISPR/Cas13a system. Gene 2024; 921:148527. [PMID: 38710293 DOI: 10.1016/j.gene.2024.148527] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2023] [Revised: 04/04/2024] [Accepted: 05/01/2024] [Indexed: 05/08/2024]
Abstract
The E6 protein is a known oncogene in cervical cancer and plays a key role in the development and progression of cervical cancer by reducing the expression level of the tumor suppressor protein P53 and ultimately leading to enhanced cell proliferation and reduced apoptosis. Therefore, antiviral agents that inhibit the expression of E6 oncoprotein are expected to be potential therapies for human cervical cancer. Here we developed CRISPR/Cas13a: crRNA dual plasmid system and demonstrated that CRISPR/Cas13a could effectively and specifically knock down human papillomavirus 18 E6 mRNA, downregulate the expression level of E6 protein, and restore the expression of the tumor suppressor gene P53 protein, thereby inhibiting the growth of cervical cancer cells and increasing their apoptosis, the E6-2, E6-3, and E6-5 groups resulted in apoptosis rates of 25.4%, 22.4%, and 22.2% in HeLa cells. Moreover, CRISPR/Cas13a enhances the proliferation inhibition and apoptosis induction of cisplatin in cervical cancer HeLa cells. The CRISPR/Cas13a system targeting HPV E6 mRNA may be a promising therapeutic approach for the treatment of human papillomavirus-associated cervical cancer.
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Affiliation(s)
- Anran Zhang
- Department of Epidemiology and Health Statistics, College of Public Health, Zhengzhou University, No. 100 Kexue Avenue, Zhengzhou, Henan 450001, People's Republic of China
| | - Xue Zheng
- Department of Epidemiology and Health Statistics, College of Public Health, Zhengzhou University, No. 100 Kexue Avenue, Zhengzhou, Henan 450001, People's Republic of China
| | - Shuaiyin Chen
- Department of Epidemiology and Health Statistics, College of Public Health, Zhengzhou University, No. 100 Kexue Avenue, Zhengzhou, Henan 450001, People's Republic of China.
| | - Guangcai Duan
- Department of Epidemiology and Health Statistics, College of Public Health, Zhengzhou University, No. 100 Kexue Avenue, Zhengzhou, Henan 450001, People's Republic of China; Henan Key Laboratory of Molecular Medicine, Zhengzhou University, No. 100 Kexue Avenue, Zhengzhou, Henan 450001, People's Republic of China.
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9
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Han J, Jang KL. All-trans retinoic acid downregulates HBx levels via E6-associated protein-mediated proteasomal degradation to suppress hepatitis B virus replication. PLoS One 2024; 19:e0305350. [PMID: 38861553 PMCID: PMC11166335 DOI: 10.1371/journal.pone.0305350] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2024] [Accepted: 05/28/2024] [Indexed: 06/13/2024] Open
Abstract
All-trans retinoic acid (ATRA), recognized as the principal and most biologically potent metabolite of vitamin A, has been identified for its inhibitory effects on hepatitis B virus (HBV) replication. Nevertheless, the underlying mechanism remains elusive. The present study reveals that ATRA induces E6-associated protein (E6AP)-mediated proteasomal degradation of HBx to suppress HBV replication in human hepatoma cells in a p53-dependent pathway. For this effect, ATRA induced promoter hypomethylation of E6AP in the presence of HBx, which resulted in the upregulation of E6AP levels in HepG2 but not in Hep3B cells, emphasizing the p53-dependent nature of this effect. As a consequence, ATRA augmented the interaction between E6AP and HBx, resulting in substantial ubiquitination of HBx and consequent reduction in HBx protein levels in both the HBx overexpression system and the in vitro HBV replication model. Additionally, the knockdown of E6AP under ATRA treatment reduced the interaction between HBx and E6AP and decreased the ubiquitin-dependent proteasomal degradation of HBx, which prompted a recovery of HBV replication in the presence of ATRA, as confirmed by increased levels of intracellular HBV proteins and secreted HBV levels. This study not only contributes to the understanding of the complex interactions between ATRA, p53, E6AP, and HBx but also provides an academic basis for the clinical employment of ATRA in the treatment of HBV infection.
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Affiliation(s)
- Jiwoo Han
- Department of Integrated Biological Science, The Graduate School, Pusan National University, Busan, Republic of Korea
| | - Kyung Lib Jang
- Department of Integrated Biological Science, The Graduate School, Pusan National University, Busan, Republic of Korea
- Department of Microbiology, College of Natural Science, Pusan National University, Busan, Republic of Korea
- Microbiological Resource Research Institute, Pusan National University, Busan, Republic of Korea
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10
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Yoshimoto M, Tokuda A, Eguchi A, Nozawa Y, Mori T, Yaginuma Y. Alterations of UHRF family Expression and was regulated by High Risk Type HPV16 in Uterine Cervical Cancer. Exp Cell Res 2024; 437:114018. [PMID: 38556072 DOI: 10.1016/j.yexcr.2024.114018] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/06/2024] [Revised: 03/20/2024] [Accepted: 03/24/2024] [Indexed: 04/02/2024]
Abstract
The altered protein expression of inverted CCAAT box-binding protein of 90 kDa/ubiquitin-like with PHD and RING finger domains 1 (ICBP90/UHRF1), and Np95-like ring finger protein (NIRF)/UHRF2, which belong to the ubiquitin-like with PHD and RING finger domains (UHRF) family, is linked to tumor malignancy and the progression of various cancers. In this study, we analyzed the UHRF family expression in cervical cancers, and it's regulation by human papillomavirus (HPV). Western blotting was performed to analyze protein expression in cervical cancer cell lines. Immunohistochemical analysis were used to investigate the expression of UHRF family and MIB-1 in cervical cancer tissues. Transfection were done for analyze the relationship between UHRF family and HPVs. We showed that NIRF expression was decreased and ICBP90 expression was increased in cervical cancers compared to normal counterparts. Western blotting also showed that NIRF expression was quite low levels, but ICBP90 was high in human cervical cancer cell lines. Interestingly, ICBP90 was up regulated by high risk type HPV16 E6 and E7, but not low-risk type HPV11. On the other hand, NIRF was down regulated by high risk type HPV16 E6 but not by E7. Low risk type HPV11 E6 did not affect the NIRF expression at all. We propose that ICBP90 overexpression, and reduced NIRF expression, found in cervical cancers, is an important event of a cervical carcinogenesis, and especially ICBP90 may offer a proliferating marker and therapeutic target for treating uterine cervical cancers.
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Affiliation(s)
- Masafumi Yoshimoto
- Department of Oncology, Graduate School of Health Sciences, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan
| | - Aoi Tokuda
- Department of Oncology, Graduate School of Health Sciences, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan
| | - Ayami Eguchi
- Department of Oncology, Graduate School of Health Sciences, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan
| | - Yoshihiro Nozawa
- Department of Pathology, Shirakawa Kosei General Hospital, Shirakawa, Japan
| | - Tsutomu Mori
- Department of Human Lifesciences, Fukushima Medical University School of Nursing, Fukushima, Japan
| | - Yuji Yaginuma
- Department of Oncology, Graduate School of Health Sciences, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan.
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11
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Zhitkevich A, Bayurova E, Avdoshina D, Zakirova N, Frolova G, Chowdhury S, Ivanov A, Gordeychuk I, Palefsky JM, Isaguliants M. HIV-1 Reverse Transcriptase Expression in HPV16-Infected Epidermoid Carcinoma Cells Alters E6 Expression and Cellular Metabolism, and Induces a Hybrid Epithelial/Mesenchymal Cell Phenotype. Viruses 2024; 16:193. [PMID: 38399969 PMCID: PMC10892743 DOI: 10.3390/v16020193] [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: 12/21/2023] [Revised: 01/22/2024] [Accepted: 01/24/2024] [Indexed: 02/25/2024] Open
Abstract
The high incidence of epithelial malignancies in HIV-1 infected individuals is associated with co-infection with oncogenic viruses, such as high-risk human papillomaviruses (HR HPVs), mostly HPV16. The molecular mechanisms underlying the HIV-1-associated increase in epithelial malignancies are not fully understood. A collaboration between HIV-1 and HR HPVs in the malignant transformation of epithelial cells has long been anticipated. Here, we delineated the effects of HIV-1 reverse transcriptase on the in vitro and in vivo properties of HPV16-infected cervical cancer cells. A human cervical carcinoma cell line infected with HPV16 (Ca Ski) was made to express HIV-1 reverse transcriptase (RT) by lentiviral transduction. The levels of the mRNA of the E6 isoforms and of the factors characteristic to the epithelial/mesenchymal transition were assessed by real-time RT-PCR. The parameters of glycolysis and mitochondrial respiration were determined using Seahorse technology. RT expressing Ca Ski subclones were assessed for the capacity to form tumors in nude mice. RT expression increased the expression of the E6*I isoform, modulated the expression of E-CADHERIN and VIMENTIN, indicating the presence of a hybrid epithelial/mesenchymal phenotype, enhanced glycolysis, and inhibited mitochondrial respiration. In addition, the expression of RT induced phenotypic alterations impacting cell motility, clonogenic activity, and the capacity of Ca Ski cells to form tumors in nude mice. These findings suggest that HIV-RT, a multifunctional protein, affects HPV16-induced oncogenesis, which is achieved through modulation of the expression of the E6 oncoprotein. These results highlight a complex interplay between HIV antigens and HPV oncoproteins potentiating the malignant transformation of epithelial cells.
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Affiliation(s)
- Alla Zhitkevich
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, 119991 Moscow, Russia; (E.B.); (D.A.); (G.F.); (I.G.)
| | - Ekaterina Bayurova
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, 119991 Moscow, Russia; (E.B.); (D.A.); (G.F.); (I.G.)
- Gamaleya National Research Center for Epidemiology and Microbiology, 123098 Moscow, Russia;
| | - Darya Avdoshina
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, 119991 Moscow, Russia; (E.B.); (D.A.); (G.F.); (I.G.)
| | - Natalia Zakirova
- Centre for Precision Genome Editing and Genetic Technologies for Biomedicine, Engelhardt Institute of Molecular Biology, 119991 Moscow, Russia;
| | - Galina Frolova
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, 119991 Moscow, Russia; (E.B.); (D.A.); (G.F.); (I.G.)
| | - Sona Chowdhury
- Division of Infectious Diseases, Department of Medicine, University of California, San Francisco, CA 94143, USA; (S.C.); (J.M.P.)
| | - Alexander Ivanov
- Gamaleya National Research Center for Epidemiology and Microbiology, 123098 Moscow, Russia;
- Centre for Precision Genome Editing and Genetic Technologies for Biomedicine, Engelhardt Institute of Molecular Biology, 119991 Moscow, Russia;
| | - Ilya Gordeychuk
- Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences, 119991 Moscow, Russia; (E.B.); (D.A.); (G.F.); (I.G.)
- Gamaleya National Research Center for Epidemiology and Microbiology, 123098 Moscow, Russia;
| | - Joel M. Palefsky
- Division of Infectious Diseases, Department of Medicine, University of California, San Francisco, CA 94143, USA; (S.C.); (J.M.P.)
| | - Maria Isaguliants
- Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, 171 77 Stockholm, Sweden
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12
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Lo Cigno I, Calati F, Girone C, Borgogna C, Venuti A, Boldorini R, Gariglio M. SIRT1 is an actionable target to restore p53 function in HPV-associated cancer therapy. Br J Cancer 2023; 129:1863-1874. [PMID: 37838812 PMCID: PMC10667542 DOI: 10.1038/s41416-023-02465-x] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/05/2023] [Revised: 09/26/2023] [Accepted: 10/04/2023] [Indexed: 10/16/2023] Open
Abstract
BACKGROUND Our aim was to evaluate the efficacy and anti-cancer action of a precision medicine approach involving a novel SIRT1-dependent pathway that, when disrupted, leads to the restoration of a functional p53 in human papillomavirus (HPV)-transformed cells. METHODS The anticancer potential of inhibiting SIRT1 was evaluated by examining the effects of the specific SIRT1 inhibitor EX527 (also known as Selisistat) or genetic silencing, either individually or in conjunction with standard chemotherapeutic agents, on a range of HPV+ cancer cells and a preclinical mouse model of HPV16-induced cancer. RESULTS We show that SIRT1 inhibition restores a transcriptionally active K382-acetylated p53 in HPV+ but not HPV- cell lines, which in turn promotes G0/G1 cell cycle arrest and inhibits clonogenicity specifically in HPV+ cells. Additionally, EX527 treatment increases the sensitivity of HPV+ cells to sublethal doses of standard genotoxic agents. The enhanced sensitivity to cisplatin as well as p53 restoration were also observed in an in vivo tumorigenicity assay using syngeneic C3.43 cells harbouring an integrated HPV16 genome, injected subcutaneously into C57BL/6J mice. CONCLUSIONS Our findings uncover an essential role of SIRT1 in HPV-driven oncogenesis, which may have direct translational implications for the treatment of this type of cancer.
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Affiliation(s)
- Irene Lo Cigno
- Virology Unit, Department of Translational Medicine, Eastern Piedmont University, Novara, Italy
| | - Federica Calati
- Virology Unit, Department of Translational Medicine, Eastern Piedmont University, Novara, Italy
| | - Carlo Girone
- Virology Unit, Department of Translational Medicine, Eastern Piedmont University, Novara, Italy
| | - Cinzia Borgogna
- Virology Unit, Department of Translational Medicine, Eastern Piedmont University, Novara, Italy
| | - Aldo Venuti
- HPV Unit, UOSD Tumor Immunology and Immunotherapy, IRCCS Regina Elena National Cancer Institute, Rome, Italy
| | - Renzo Boldorini
- Pathology Unit, Department of Health Sciences, Eastern Piedmont University, Novara, Italy
| | - Marisa Gariglio
- Virology Unit, Department of Translational Medicine, Eastern Piedmont University, Novara, Italy.
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13
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Chaudhary P, Proulx J, Park IW. Ubiquitin-protein ligase E3A (UBE3A) mediation of viral infection and human diseases. Virus Res 2023; 335:199191. [PMID: 37541588 PMCID: PMC10430597 DOI: 10.1016/j.virusres.2023.199191] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/05/2023] [Revised: 08/01/2023] [Accepted: 08/02/2023] [Indexed: 08/06/2023]
Abstract
The Ubiquitin-protein ligase E3A, UBE3A, also known as E6-associated protein (E6-AP), is known to play an essential role in regulating the degradation of various proteins by transferring Ub from E2 Ub conjugating enzymes to the substrate proteins. Several studies indicate that UBE3A regulates the stabilities of key viral proteins in the virus-infected cells and, thereby, the infected virus-mediated diseases, even if it were reported that UBE3A participates in non-viral-related human diseases. Furthermore, mutations such as deletions and duplications in the maternally inherited gene in the brain cause human neurodevelopmental disorders such as Angelman syndrome (AS) and autism. It is also known that UBE3A functions as a transcriptional coactivator for the expression of steroid hormone receptors. These reports establish that UBE3A is distinguished by its multitudinous functions that are paramount to viral pathology and human diseases. This review is focused on molecular mechanisms for such intensive participation of UBE3A in disease formation and virus regulation.
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Affiliation(s)
- Pankaj Chaudhary
- Department of Microbiology, Immunology and Genetics, School of Biomedical Sciences, University of North Texas Health Science Center, Fort Worth, TX 76107, United States.
| | - Jessica Proulx
- Department of Microbiology, Immunology and Genetics, School of Biomedical Sciences, University of North Texas Health Science Center, Fort Worth, TX 76107, United States
| | - In-Woo Park
- Department of Microbiology, Immunology and Genetics, School of Biomedical Sciences, University of North Texas Health Science Center, Fort Worth, TX 76107, United States.
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14
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Luo Y, Yang Y, Yang C, Li C, Hu R, Geng W, Kang X, Lin H. UBE3A and MCM6 synergistically regulate the proliferation and migration of lung adenocarcinoma cells. FEBS Open Bio 2023; 13:1756-1771. [PMID: 37454373 PMCID: PMC10476561 DOI: 10.1002/2211-5463.13675] [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: 02/07/2023] [Revised: 05/16/2023] [Accepted: 07/13/2023] [Indexed: 07/18/2023] Open
Abstract
Lung cancer is a leading cause of mortality worldwide and shows substantial clinical and biomolecular heterogeneity. Currently, specific therapeutic strategies are lacking, so effective drug targets are urgently needed. E6AP/UBE3A is a multifaceted ubiquitin ligase that controls various signaling pathways implicated in neurological diseases and various cancers; however, its role in lung cancer is incompletely understood. Here, MCM6 was identified as an interacting partner of E6AP using the yeast two-hybrid assay. MCM2 and MCM4 were then shown to interact with E6AP. E6AP knockout enhanced the ubiquitination of MCM2/4/6, suggesting that E6AP was not the E3 ubiquitin ligase for these three MCM proteins. Ablation of E6AP inhibited proliferation and migration, but had no significant effect on apoptosis in A549 and H1975 cells, and proliferation and migration inhibition was also observed in MCM6 knockdown cells. Furthermore, ablation of MCM6 and E6AP synergistically suppressed the proliferation and migration of A549 and H1975 cells. To verify the above findings in vivo, we established tumor models in nude mice and identified that the tumorigenicity of human lung adenocarcinoma (LUAD) cells was synergistically regulated by MCM6 and E6AP. Moreover, the expression levels of MCM6 and E6AP were higher in LUAD tissues than in adjacent tissues. Furthermore, the expression levels of MCM6 and E6AP were positively correlated in human LUAD samples. Thus, our study suggests that the interaction of E6AP and MCM proteins plays an important role in the progression of LUAD, which might offer potential therapeutic targets for cancer treatment.
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Affiliation(s)
- Yanyan Luo
- Department of Pain, Wenzhou Key Laboratory of Perioperative MedicineThe First Affiliated Hospital of Wenzhou Medical UniversityChina
- State Key Laboratory of Molecular Biology, Shanghai Institute of Biochemistry and Cell BiologyCenter for Excellence in Molecular Cell Science, Chinese Academy of SciencesShanghaiChina
| | - Yun Yang
- State Key Laboratory of Molecular Biology, Shanghai Institute of Biochemistry and Cell BiologyCenter for Excellence in Molecular Cell Science, Chinese Academy of SciencesShanghaiChina
- School of MedicineGuizhou UniversityGuiyangChina
| | - Cong Yang
- Cancer Center, School of Medicine, Shanghai Tenth People's HospitalfTongji UniversityShanghaiChina
| | - Chuanyin Li
- Cancer Center, School of Medicine, Shanghai Tenth People's HospitalfTongji UniversityShanghaiChina
| | - Ronggui Hu
- Department of Pain, Wenzhou Key Laboratory of Perioperative MedicineThe First Affiliated Hospital of Wenzhou Medical UniversityChina
- State Key Laboratory of Molecular Biology, Shanghai Institute of Biochemistry and Cell BiologyCenter for Excellence in Molecular Cell Science, Chinese Academy of SciencesShanghaiChina
| | - Wujun Geng
- Department of Pain, Wenzhou Key Laboratory of Perioperative MedicineThe First Affiliated Hospital of Wenzhou Medical UniversityChina
| | - Xianhui Kang
- Department of Pain, Wenzhou Key Laboratory of Perioperative MedicineThe First Affiliated Hospital of Wenzhou Medical UniversityChina
- Department of Anesthesiology, The First Affiliated HospitalZhejiang University School of MedicineHangzhouChina
| | - Hai Lin
- Department of Pain, Wenzhou Key Laboratory of Perioperative MedicineThe First Affiliated Hospital of Wenzhou Medical UniversityChina
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15
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Mink JN, Khalmurzaev O, Pryalukhin A, Hölters S, Geppert C, Lohse S, Bende K, Lobo J, Henrique R, Loertzer H, Steffens J, Jerónimo C, Wunderlich H, Heinzelbecker J, Bohle R, Stöckle M, Matveev V, Hartmann A, Junker K. [Importance of HPV status and p16 for the prognosis of penile carcinoma]. Aktuelle Urol 2023; 54:274-284. [PMID: 37541236 DOI: 10.1055/a-2104-1270] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 08/06/2023]
Abstract
BACKGROUND Penile cancer is a rare but often lethal tumour disease, especially in the metastatic stage. Most data on prognostic factors for penile cancer are based on small patient cohorts, and even meta-analyses are mostly limited in terms of patient numbers. There is a lack of sufficient parameters to predict the metastatic risk of these tumours. Furthermore, the role of the HPV status for the prognosis, and, in this regard, of p16INK4a is still unclear. MATERIAL AND METHODS In this study, 236 patients from an international multicentre cohort were analysed with regard to histological subtypes, HPV and p16 status, and other clinical parameters. The HPV status was only graded as HPV-positive if HPV was detected by PCR and the p16 status defined by immunochemistry was positive. The statistical analysis was carried out using the Kaplan-Meier method as well as the log-rank test and a univariable and multivariable analysis using the Cox regression model. RESULTS A positive HPV status was not a significant parameter for either metastasis-free (MFS), tumour-specific (CSS) or overall survival (OS). p16-positive tumours showed a significantly better MFS (p=0.026), which was also confirmed in the subgroup analysis of HPV-negative tumours (p=0.037) without differences in CSS or OS. In the usual type, there was also a trend towards an improved MFS, but without statistical significance (p=0.070). p16-positive tumours were associated with a highly significantly better MFS (hazard ratio 0.3; p=0.004) in the multivariable Cox regression, while patients with a pT1b tumour stage or advanced lymph node metastasis showed a significantly worse survival. In the multivariable analysis of HPV-negative tumours, p16 status was also confirmed as an independent predictor of MFS (Hazard ratio 0.2; p=0.007). CONCLUSION HPV status alone seems to be lacking prognostic relevance. In contrast, p16 status was confirmed as an independent prognostic factor. Thus, the expression of p16INK4a is associated with a significantly better MFS. Especially in HPV-negative tumours, the p16 status should be evaluated with regard to the prognostic value and thus also with a view to the treatment decision.
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Affiliation(s)
- Jan Niklas Mink
- Klinik für Urologie und Kinderurologie, Universitätsklinikum des Saarlandes und Medizinische Fakultät der Universität des Saarlandes, Homburg, Deutschland
| | - Oybek Khalmurzaev
- Klinik für Urologie und Kinderurologie, Universitätsklinikum des Saarlandes und Medizinische Fakultät der Universität des Saarlandes, Homburg, Deutschland
- Abteilung für Urologie, N N Blokhin NMRCO, Moskva, Russian Federation
| | - Alexey Pryalukhin
- Abteilung für Pathologie, Universitätsklinikum des Saarlandes und Medizinische Fakultät der Universität des Saarlandes, Homburg, Deutschland
- Institut für Pathologie, Universitätsklinikum Bonn, Bonn, Deutschland
| | - Sebastian Hölters
- Klinik für Urologie und Kinderurologie, Universitätsklinikum des Saarlandes und Medizinische Fakultät der Universität des Saarlandes, Homburg, Deutschland
| | - Carol Geppert
- Institut für Pathologie, Universitätsklinikum Erlangen, Erlangen, Deutschland
| | - Stefan Lohse
- Abteilung für Virologie, Universitätsklinikum des Saarlandes und Medizinische Fakultät der Universität des Saarlandes, Homburg, Deutschland
| | - Kristof Bende
- Institut für Pathologie, Universitätsklinikum Erlangen, Erlangen, Deutschland
| | - João Lobo
- Abteilung für Pathologie, IPO-PORTO, Porto, Portugal
| | - Rui Henrique
- Abteilung für Pathologie, IPO-PORTO, Porto, Portugal
| | - Hagen Loertzer
- Abteilung für Urologie und Kinderurologie, Westpfalz Klinikum Gmbh, Kaiserslautern, Deutschland
| | - Joachim Steffens
- Klinik für Urologie und Kinderurologie, St-Antonius-Hospital gGmbH, Eschweiler, Deutschland
| | - Carmen Jerónimo
- Abteilung für Pathologie, IPO-PORTO, Porto, Portugal
- Portuguese Oncology Institute of Porto, Research Center (CI-IPOP), Porto
| | - Heiko Wunderlich
- Abteilung für Urologie und Kinderurologie, St. Georg Klinikum Eisenach, Eisenach, Deutschland
| | - Julia Heinzelbecker
- Klinik für Urologie und Kinderurologie, Universitätsklinikum des Saarlandes und Medizinische Fakultät der Universität des Saarlandes, Homburg, Deutschland
| | - Rainer Bohle
- Abteilung für Pathologie, Universitätsklinikum des Saarlandes und Medizinische Fakultät der Universität des Saarlandes, Homburg, Deutschland
| | - Michael Stöckle
- Klinik für Urologie und Kinderurologie, Universitätsklinikum des Saarlandes und Medizinische Fakultät der Universität des Saarlandes, Homburg, Deutschland
| | - Vsevolod Matveev
- Abteilung für Urologie, N N Blokhin NMRCO, Moskva, Russian Federation
| | - Arndt Hartmann
- Institut für Pathologie, Universitätsklinikum Erlangen, Erlangen, Deutschland
| | - Kerstin Junker
- Klinik für Urologie und Kinderurologie, Universitätsklinikum des Saarlandes und Medizinische Fakultät der Universität des Saarlandes, Homburg, Deutschland
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Paulis A, Tramontano E. Unlocking STING as a Therapeutic Antiviral Strategy. Int J Mol Sci 2023; 24:ijms24087448. [PMID: 37108610 PMCID: PMC10138487 DOI: 10.3390/ijms24087448] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/11/2023] [Revised: 04/06/2023] [Accepted: 04/11/2023] [Indexed: 04/29/2023] Open
Abstract
Invading pathogens have developed weapons that subvert physiological conditions to weaken the host and permit the spread of infection. Cells, on their side, have thus developed countermeasures to maintain cellular physiology and counteract pathogenesis. The cyclic GMP-AMP (cGAMP) synthase (cGAS) is a pattern recognition receptor that recognizes viral DNA present in the cytosol, activating the stimulator of interferon genes (STING) protein and leading to the production of type I interferons (IFN-I). Given its role in innate immunity activation, STING is considered an interesting and innovative target for the development of broad-spectrum antivirals. In this review, we discuss the function of STING; its modulation by the cellular stimuli; the molecular mechanisms developed by viruses, through which they escape this defense system; and the therapeutical strategies that have been developed to date to inhibit viral replication restoring STING functionality.
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Affiliation(s)
- Annalaura Paulis
- Department of Life and Environmental Sciences, Università Degli Studi di Cagliari, 09124 Cagliari, Italy
| | - Enzo Tramontano
- Department of Life and Environmental Sciences, Università Degli Studi di Cagliari, 09124 Cagliari, Italy
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17
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Zhai F, Wang J, Yang W, Ye M, Jin X. The E3 Ligases in Cervical Cancer and Endometrial Cancer. Cancers (Basel) 2022; 14:5354. [PMID: 36358773 PMCID: PMC9658772 DOI: 10.3390/cancers14215354] [Citation(s) in RCA: 11] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2022] [Revised: 10/25/2022] [Accepted: 10/27/2022] [Indexed: 07/28/2023] Open
Abstract
Endometrial (EC) and cervical (CC) cancers are the most prevalent malignancies of the female reproductive system. There is a global trend towards increasing incidence and mortality, with a decreasing age trend. E3 ligases label substrates with ubiquitin to regulate their activity and stability and are involved in various cellular functions. Studies have confirmed abnormal expression or mutations of E3 ligases in EC and CC, indicating their vital roles in the occurrence and progression of EC and CC. This paper provides an overview of the E3 ligases implicated in EC and CC and discusses their underlying mechanism. In addition, this review provides research advances in the target of ubiquitination processes in EC and CC.
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Affiliation(s)
- Fengguang Zhai
- The Affiliated Hospital of Medical School, Ningbo University, Ningbo 315020, China
- Department of Biochemistry and Molecular Biology, Zhejiang Key Laboratory of Pathophysiology, Medical School of Ningbo University, Ningbo 315211, China
| | - Jie Wang
- The Affiliated Hospital of Medical School, Ningbo University, Ningbo 315020, China
- Department of Biochemistry and Molecular Biology, Zhejiang Key Laboratory of Pathophysiology, Medical School of Ningbo University, Ningbo 315211, China
| | - Weili Yang
- Department of Gynecology, The Affiliated People’s Hospital of Ningbo University, Ningbo 315040, China
| | - Meng Ye
- The Affiliated Hospital of Medical School, Ningbo University, Ningbo 315020, China
- Department of Biochemistry and Molecular Biology, Zhejiang Key Laboratory of Pathophysiology, Medical School of Ningbo University, Ningbo 315211, China
| | - Xiaofeng Jin
- The Affiliated Hospital of Medical School, Ningbo University, Ningbo 315020, China
- Department of Biochemistry and Molecular Biology, Zhejiang Key Laboratory of Pathophysiology, Medical School of Ningbo University, Ningbo 315211, China
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18
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Tumor Suppressor p53 Inhibits Hepatitis B Virus Replication by Downregulating HBx via E6AP-Mediated Proteasomal Degradation in Human Hepatocellular Carcinoma Cell Lines. Viruses 2022; 14:v14102313. [PMID: 36298868 PMCID: PMC9609658 DOI: 10.3390/v14102313] [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: 09/30/2022] [Revised: 10/18/2022] [Accepted: 10/19/2022] [Indexed: 11/06/2022] Open
Abstract
HBx, a multifunctional regulatory protein, plays an essential role in the replication and pathogenesis of the hepatitis B virus (HBV). In this study, we found that in human hepatoma cells, the tumor suppressor p53 downregulates HBx via ubiquitin-dependent proteasomal degradation. p53 transcriptional activity that results from HBV infection was not essential for this effect. This was shown by treatment with a potent p53 inhibitor, pifithrin-α. Instead, we found that p53 facilitated the binding of E6-associated protein (E6AP), which is an E3 ligase, to HBx and induced E6AP-mediated HBx ubiquitination in a ternary complex of p53, E6AP, and HBx. The ability of p53 to induce E6AP-mediated downregulation of HBx and inhibit HBV replication was demonstrated in an in vitro HBV infection system. This study may provide insights into the regulation of HBx and HBV replication, especially with respect to p53 status, which may also help in understanding HBV-associated tumorigenesis in patients.
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19
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Lim J, Frecot DI, Stubenrauch F, Iftner T, Simon C. Cottontail rabbit papillomavirus E6 proteins: Interaction with MAML1 and modulation of the Notch signaling pathway. Virology 2022; 576:52-60. [PMID: 36155393 DOI: 10.1016/j.virol.2022.08.014] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/23/2022] [Revised: 08/22/2022] [Accepted: 08/22/2022] [Indexed: 12/01/2022]
Abstract
Animal models are necessary to study how cutaneous human papillomaviruses (HPVs) are associated with carcinogenesis. The cottontail rabbit papillomavirus (CRPV) induces papilloma in the -cutaneous skin of rabbits and serves as an established animal model for HPVlinked carcinogenesis where viral E6 proteins play crucial roles. Several studies have reported the dysregulation of the Notch signaling pathway by cutaneous beta HPV, bovine PV and mouse PV E6 via their association with Mastermind-like 1 protein (MAML1), thus interfering with cell proliferation and differentiation. However, the CRPV E6 gene encodes an elongated E6 protein (long E6, LE6) and an N-terminally truncated product (short E6, SE6) making it unique from other E6 proteins. Here, we describe the interaction between both CRPV E6 proteins and MAML1 and their ability to downregulate the Notch signaling pathway which could be a way CRPV infection induces carcinogenesis similar to beta HPV.
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Affiliation(s)
- JiaWen Lim
- Institute of Medical Virology and Epidemiology of Viral Diseases, University Hospital Tuebingen, Tuebingen, Germany
| | - Desiree Isabella Frecot
- Institute of Medical Virology and Epidemiology of Viral Diseases, University Hospital Tuebingen, Tuebingen, Germany
| | - Frank Stubenrauch
- Institute of Medical Virology and Epidemiology of Viral Diseases, University Hospital Tuebingen, Tuebingen, Germany
| | - Thomas Iftner
- Institute of Medical Virology and Epidemiology of Viral Diseases, University Hospital Tuebingen, Tuebingen, Germany.
| | - Claudia Simon
- Institute of Medical Virology and Epidemiology of Viral Diseases, University Hospital Tuebingen, Tuebingen, Germany.
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20
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Park JM, Yoon H, Jeong Y, Jang KL. Tumor suppressor p53 inhibits hepatitis C virus replication by inducing E6AP-mediated proteasomal degradation of the viral core protein. FEBS Lett 2022; 596:2525-2537. [PMID: 35918185 DOI: 10.1002/1873-3468.14461] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/05/2022] [Revised: 06/24/2022] [Accepted: 07/09/2022] [Indexed: 11/08/2022]
Abstract
The tumor suppressor p53 has been implicated in the host defense system against hepatitis C virus (HCV) infection, although the detailed mechanism remains unknown. Here, we found that p53 inhibits HCV replication by downregulating HCV Core protein levels in human hepatoma cells. For this effect, p53 potentiated the role of E6-associated protein (E6AP) as an E3 ligase to induce ubiquitination and proteasomal degradation of HCV Core. Specifically, p53 facilitated the binding of E6AP to HCV Core through direct interactions with the two proteins. In addition, E6AP failed to induce ubiquitination of HCV Core in the absence of p53, suggesting that p53 increases the E3 ligase activity of E6AP in a triple complex consisting of p53, E6AP, and HCV Core.
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Affiliation(s)
- Ji-Min Park
- Department of Microbiology, College of Natural Science, Pusan National University, Busan, 46241, Republic of Korea
| | - Hyunyoung Yoon
- Department of Microbiology, College of Natural Science, Pusan National University, Busan, 46241, Republic of Korea
| | - Yuna Jeong
- Department of Microbiology, College of Natural Science, Pusan National University, Busan, 46241, Republic of Korea
| | - Kyung Lib Jang
- Department of Microbiology, College of Natural Science, Pusan National University, Busan, 46241, Republic of Korea.,Microbiological Resource Research Institute, Pusan National University, Busan, 46241, Republic of Korea
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21
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Makgoo L, Mosebi S, Mbita Z. Molecular Mechanisms of HIV Protease Inhibitors Against HPV-Associated Cervical Cancer: Restoration of TP53 Tumour Suppressor Activities. Front Mol Biosci 2022; 9:875208. [PMID: 35620479 PMCID: PMC9127998 DOI: 10.3389/fmolb.2022.875208] [Citation(s) in RCA: 9] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/13/2022] [Accepted: 04/12/2022] [Indexed: 12/24/2022] Open
Abstract
Cervical cancer is a Human Papilloma virus-related disease, which is on the rise in a number of countries, globally. Two essential oncogenes, E6 and E7, drive cell transformation and cancer development. These two oncoproteins target two of the most important tumour suppressors, p53 and pRB, for degradation through the ubiquitin ligase pathway, thus, blocking apoptosis activation and deregulation of cell cycle. This pathway can be exploited for anticancer therapeutic interventions, and Human Immunodeficiency Virus Protease Inhibitors (HIV-PIs) have attracted a lot of attention for this anticancer drug development. HIV-PIs have proven effective in treating HPV-positive cervical cancers and shown to restore impaired or deregulated p53 in HPV-associated cervical cancers by inhibiting the 26S proteasome. This review will evaluate the role players, such as HPV oncoproteins involved cervical cancer development and how they are targeted in HIV protease inhibitors-induced p53 restoration in cervical cancer. This review also covers the therapeutic potential of HIV protease inhibitors and molecular mechanisms behind the HIV protease inhibitors-induced p53-dependent anticancer activities against cervical cancer.
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Affiliation(s)
- Lilian Makgoo
- Department of Biochemistry, Microbiology and Biotechnology, University of Limpopo, Sovenga, South Africa
| | - Salerwe Mosebi
- Department of Life and Consumer Sciences, University of South Africa, Florida, South Africa
| | - Zukile Mbita
- Department of Biochemistry, Microbiology and Biotechnology, University of Limpopo, Sovenga, South Africa
- *Correspondence: Zukile Mbita,
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22
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An enhanced triple fluorescence flow-cytometry-based assay shows differential activation of the Notch signaling pathway by human papillomavirus E6 proteins. Sci Rep 2022; 12:3000. [PMID: 35194094 PMCID: PMC8863805 DOI: 10.1038/s41598-022-06922-0] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2021] [Accepted: 01/27/2022] [Indexed: 01/07/2023] Open
Abstract
Human papillomaviruses are DNA tumor viruses. A persistent infection with high-risk HPV types is the necessary risk factor for the development of anogenital carcinoma. The E6 protein is a viral oncoprotein that directly interacts with different cellular regulatory proteins mainly affecting the cell cycle, cellular differentiation and polarization of epithelial cells. In dependency of the phylogenetic classification of HPV different interaction partners of E6 have been described. The Notch pathway seems to be one common target of HPV, which can be up or down regulated by different E6 proteins. Our novel triple fluorescence flow-cytometry-based assay allows a semi-quantitative comparison of the E6 proteins´ effect on the Notch pathway using a Notch-responsive reporter plasmid. As a result, all E6 proteins of beta-HPV repressed the Notch reporter expression, of which HPV38 E6 showed the greatest repression potential. In contrast, alpha-HPV E6 of HPV16, activates the reporter expression most significantly, whereas E6 of HPV31 and low-risk HPV6b showed significant activation only in a p53-null cell line. Interestingly, HPV18 E6, with the second highest carcinogenic risk, shows no effect. This high divergence within different genus of HPV is important for targeting the Notch pathway regarding a potential HPV therapy.
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Soh SM, Kim YJ, Kim HH, Lee HR. Modulation of Ubiquitin Signaling in Innate Immune Response by Herpesviruses. Int J Mol Sci 2022; 23:ijms23010492. [PMID: 35008917 PMCID: PMC8745310 DOI: 10.3390/ijms23010492] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/20/2021] [Revised: 12/30/2021] [Accepted: 12/31/2021] [Indexed: 12/16/2022] Open
Abstract
The ubiquitin proteasome system (UPS) is a protein degradation machinery that is crucial for cellular homeostasis in eukaryotes. Therefore, it is not surprising that the UPS coordinates almost all host cellular processes, including host-pathogen interactions. This protein degradation machinery acts predominantly by tagging substrate proteins designated for degradation with a ubiquitin molecule. These ubiquitin tags have been involved at various steps of the innate immune response. Hence, herpesviruses have evolved ways to antagonize the host defense mechanisms by targeting UPS components such as ubiquitin E3 ligases and deubiquitinases (DUBs) that establish a productive infection. This review delineates how herpesviruses usurp the critical roles of ubiquitin E3 ligases and DUBs in innate immune response to escape host-antiviral immune response, with particular focus on retinoic acid-inducible gene I (RIG-I)-like receptors (RLR), cyclic-GMP-AMP (cGAMP) synthase (cGAS), stimulator of interferon (IFN) genes (STING) pathways, and inflammasome signaling.
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Affiliation(s)
- Sandrine-M. Soh
- Department of Biotechnology and Bioinformatics, College of Science and Technology, Korea University, Sejong 30019, Korea; (S.-M.S.); (Y.-J.K.); (H.-H.K.)
| | - Yeong-Jun Kim
- Department of Biotechnology and Bioinformatics, College of Science and Technology, Korea University, Sejong 30019, Korea; (S.-M.S.); (Y.-J.K.); (H.-H.K.)
| | - Hong-Hee Kim
- Department of Biotechnology and Bioinformatics, College of Science and Technology, Korea University, Sejong 30019, Korea; (S.-M.S.); (Y.-J.K.); (H.-H.K.)
| | - Hye-Ra Lee
- Department of Biotechnology and Bioinformatics, College of Science and Technology, Korea University, Sejong 30019, Korea; (S.-M.S.); (Y.-J.K.); (H.-H.K.)
- Department of Laboratory Medicine, College of Medicine, Korea University, Seoul 136-701, Korea
- Correspondence: ; Tel.: +82-44-860-1831
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Wang Y, Xie Y, Sun B, Guo Y, Song L, Mohammednur DE, Zhao C. The degradation of Rap1GAP via E6AP-mediated ubiquitin-proteasome pathway is associated with HPV16/18-infection in cervical cancer cells. Infect Agent Cancer 2021; 16:71. [PMID: 34952616 PMCID: PMC8710002 DOI: 10.1186/s13027-021-00409-9] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/05/2020] [Accepted: 12/03/2021] [Indexed: 11/10/2022] Open
Abstract
BACKGROUND Cervical cancers are closely associated with persistent high-risk human papillomaviruses (HR HPV) infection. The main mechanism involves the targeting of tumor suppressors, such as p53 and pRB, for degradation by HR HPV-encoded oncoproteins, thereby leading to tumorigenesis. Rap1GAP, a tumor suppressor gene, is down-regulated in many cancers. Previous studies have revealed that down-regulation of Rap1GAP is correlated with HPV16/18 infection in cervical cancer. However, the molecular mechanism remains unclear. In this study, we aimed to address the degradation pathway of Rap1GAP in HPV-positive cervical cancer cells. METHODS HPV-positive (HeLa and SiHa) and negative (C33A) cervical cancer cells were used to analyze the pathways of Rap1GAP degradation. MG132 (carbobenzoxy-leucyl-leucyl-leucine) was used to inhibit protein degradation by proteasome. Co-immunoprecipitation (co-IP) was used to detect the interaction between Rap1GAP and E6AP. siRNA for E6AP was used to silence the expression of E6AP. Rapamycin was used to induce cell autophagy. Western blotting was used to check the levels of proteins. RESULTS Following treatment with MG132, the levels of Rap1GAP were increased in the HR HPV-positive HeLa and SiHa cells, but not in the HPV-negative C33A cells. Co-immunoprecipitation assay revealed ubiquitinated Rap1GAP protein in HeLa and SiHa cells, but not in C33A cells. E6-associated protein (E6AP) mediated the ubiquitination of Rap1GAP by binding to it in HeLa and SiHa cells, but not in C33A cells. However, the levels of Rap1GAP were decreased in HeLa and SiHa cells after knocking down E6AP by siRNA. Silencing of E6AP did not affect the levels of Rap1GAP in C33A cells. Autophagy marker p62 was decreased and LC3 II/LC3 I was increased after knocking down E6AP in HeLa cells, but not in C33A cells. The levels of Rap1GAP were decreased after treating the cells with rapamycin to induce cell autophagy in HeLa and C33A cells. CONCLUSION Rap1GAP may be degraded by autophagy in cervical cancer cells, but HPV infection can switch the degradation pathway from autophagy to E6AP-mediated ubiquitin-proteasome degradation. E6AP may be a key component of the switch.
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Affiliation(s)
- Yinghui Wang
- College of Laboratory Medicine, Dalian Medical University, 9 West Section, Lvshun South Road, Dalian, Liaoning, China
- Liaoning Provincial Center for Disease Control and Prevention, Shenyang, China
| | - Yihang Xie
- College of Laboratory Medicine, Dalian Medical University, 9 West Section, Lvshun South Road, Dalian, Liaoning, China
| | - Boxuan Sun
- College of Laboratory Medicine, Dalian Medical University, 9 West Section, Lvshun South Road, Dalian, Liaoning, China
| | - Yuwei Guo
- College of Laboratory Medicine, Dalian Medical University, 9 West Section, Lvshun South Road, Dalian, Liaoning, China
| | - Ling Song
- College of Laboratory Medicine, Dalian Medical University, 9 West Section, Lvshun South Road, Dalian, Liaoning, China
- Foruth Teaching Hospital, Tianjin University of Traditional Chinese Medicine, Tianjin, China
| | - Dawit Eman Mohammednur
- College of Laboratory Medicine, Dalian Medical University, 9 West Section, Lvshun South Road, Dalian, Liaoning, China
| | - Chunyan Zhao
- College of Laboratory Medicine, Dalian Medical University, 9 West Section, Lvshun South Road, Dalian, Liaoning, China.
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Cruz-Gregorio A, Aranda-Rivera AK, Pedraza-Chaverri J. Nuclear factor erythroid 2-related factor 2 in human papillomavirus-related cancers. Rev Med Virol 2021; 32:e2308. [PMID: 34694662 DOI: 10.1002/rmv.2308] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/21/2021] [Revised: 10/06/2021] [Accepted: 10/11/2021] [Indexed: 01/04/2023]
Abstract
High-risk human papillomavirus (HR-HPV) infection is a necessary cause for the development of cervical cancer. Moreover, HR-HPV is also associated with cancers in the anus, vagina, vulva, penis and oropharynx. HR-HPVs target and modify the function of different cell biomolecules, such as glucose, amino acids, lipids and transcription factors (TF), such as p53, nuclear factor erythroid 2-related factor 2 (Nrf2), among others. The latter is a master TF that maintains redox homeostasis. Nrf2 also induces the transcription of genes associated with cell detoxification. Since both processes are critical for cell physiology, Nrf2 deregulation is associated with cancer development. Nrf2 is a crucial molecule in HPV-related cancer development but underexplored. Moreover, Nrf2 activation is also associated with resistance to chemotherapy and radiotherapy in these cancers. This review focusses on the importance of Nrf2 during HPV-related cancer development, resistance to therapy and potential therapies associated with Nrf2 as a molecular target.
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Affiliation(s)
- Alfredo Cruz-Gregorio
- Departmento de Biología, Facultad de Química, Universidad Nacional Autónoma de México, Ciudad de Mexico, Mexico
| | - Ana Karina Aranda-Rivera
- Departmento de Biología, Facultad de Química, Universidad Nacional Autónoma de México, Ciudad de Mexico, Mexico
| | - José Pedraza-Chaverri
- Departmento de Biología, Facultad de Química, Universidad Nacional Autónoma de México, Ciudad de Mexico, Mexico
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Mathieu NA, Levin RH, Spratt DE. Exploring the Roles of HERC2 and the NEDD4L HECT E3 Ubiquitin Ligase Subfamily in p53 Signaling and the DNA Damage Response. Front Oncol 2021; 11:659049. [PMID: 33869064 PMCID: PMC8044464 DOI: 10.3389/fonc.2021.659049] [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: 01/26/2021] [Accepted: 03/16/2021] [Indexed: 12/27/2022] Open
Abstract
Cellular homeostasis is governed by the precise expression of genes that control the translation, localization, and termination of proteins. Oftentimes, environmental and biological factors can introduce mutations into the genetic framework of cells during their growth and division, and these genetic abnormalities can result in malignant transformations caused by protein malfunction. For example, p53 is a prominent tumor suppressor protein that is capable of undergoing more than 300 posttranslational modifications (PTMs) and is involved with controlling apoptotic signaling, transcription, and the DNA damage response (DDR). In this review, we focus on the molecular mechanisms and interactions that occur between p53, the HECT E3 ubiquitin ligases WWP1, SMURF1, HECW1 and HERC2, and other oncogenic proteins in the cell to explore how irregular HECT-p53 interactions can induce tumorigenesis.
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Affiliation(s)
- Nicholas A Mathieu
- Gustaf H. Carlson School of Chemistry and Biochemistry, Clark University, Worcester, MA, United States
| | - Rafael H Levin
- Gustaf H. Carlson School of Chemistry and Biochemistry, Clark University, Worcester, MA, United States
| | - Donald E Spratt
- Gustaf H. Carlson School of Chemistry and Biochemistry, Clark University, Worcester, MA, United States
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Park S, Auyeung A, Lee DL, Lambert PF, Carchman EH, Sherer NM. HIV-1 Protease Inhibitors Slow HPV16-Driven Cell Proliferation through Targeted Depletion of Viral E6 and E7 Oncoproteins. Cancers (Basel) 2021; 13:949. [PMID: 33668328 PMCID: PMC7956332 DOI: 10.3390/cancers13050949] [Citation(s) in RCA: 20] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/28/2020] [Revised: 02/09/2021] [Accepted: 02/20/2021] [Indexed: 02/05/2023] Open
Abstract
High-risk human papillomavirus strain 16 (HPV16) causes oral and anogenital cancers through the activities of two viral oncoproteins, E6 and E7, that dysregulate the host p53 and pRb tumor suppressor pathways, respectively. The maintenance of HPV16-positive cancers requires constitutive expression of E6 and E7. Therefore, inactivating these proteins could provide the basis for an anticancer therapy. Herein we demonstrate that a subset of aspartyl protease inhibitor drugs currently used to treat HIV/AIDS cause marked reductions in HPV16 E6 and E7 protein levels using two independent cell culture models: HPV16-transformed CaSki cervical cancer cells and NIKS16 organotypic raft cultures (a 3-D HPV16-positive model of epithelial pre-cancer). Treatment of CaSki cells with some (lopinavir, ritonavir, nelfinavir, and saquinavir) but not other (indinavir and atazanavir) protease inhibitors reduced E6 and E7 protein levels, correlating with increased p53 protein levels and decreased cell viability. Long-term (>7 day) treatment of HPV16-positive NIKS16 raft cultures with saquinavir caused epithelial atrophy with no discernible effects on HPV-negative rafts, demonstrating selectivity. Saquinavir also reduced HPV16's effects on markers of the cellular autophagy pathway in NIKS16 rafts, a hallmark of HPV-driven pre-cancers. Taken together, these data suggest HIV-1 protease inhibitors be studied further in the context of treating or preventing HPV16-positive cancers.
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Affiliation(s)
- Soyeong Park
- McArdle Laboratory for Cancer Research, Deptartment of Oncology, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA; (S.P.); (D.L.L.); (P.F.L.)
- Institute for Molecular Virology, University of Wisconsin-Madison, Madison, WI 53706, USA
- Carbone Cancer Center, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA; (A.A.); (E.H.C.)
| | - Andrew Auyeung
- Carbone Cancer Center, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA; (A.A.); (E.H.C.)
- Department of Surgery, University of Wisconsin School of Medicine and Public Health, Madison, WI 53792, USA
| | - Denis L. Lee
- McArdle Laboratory for Cancer Research, Deptartment of Oncology, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA; (S.P.); (D.L.L.); (P.F.L.)
- Carbone Cancer Center, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA; (A.A.); (E.H.C.)
| | - Paul F. Lambert
- McArdle Laboratory for Cancer Research, Deptartment of Oncology, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA; (S.P.); (D.L.L.); (P.F.L.)
- Carbone Cancer Center, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA; (A.A.); (E.H.C.)
| | - Evie H. Carchman
- Carbone Cancer Center, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA; (A.A.); (E.H.C.)
- Department of Surgery, University of Wisconsin School of Medicine and Public Health, Madison, WI 53792, USA
| | - Nathan M. Sherer
- McArdle Laboratory for Cancer Research, Deptartment of Oncology, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA; (S.P.); (D.L.L.); (P.F.L.)
- Institute for Molecular Virology, University of Wisconsin-Madison, Madison, WI 53706, USA
- Carbone Cancer Center, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA; (A.A.); (E.H.C.)
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Notch Signaling and Human Papillomavirus-Associated Oral Tumorigenesis. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2021; 1287:105-122. [PMID: 33034029 DOI: 10.1007/978-3-030-55031-8_8] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
Abstract
The NOTCH pathway is critical for the development of many cell types including the squamous epithelium lining of cutaneous and mucosal surfaces. In genetically engineered mouse models, Notch1 acts as one of the first steps to commit basal keratinocytes to terminally differentiate. Similarly, in human head and neck squamous cell cancers (HNSCCs), NOTCH1 is often lost consistent with its essential tumor-suppressive role for initiating keratinocyte differentiation. However, constitutive NOTCH1 activity in the epithelium results in expansion of the spinous keratinocyte layers and impaired terminal differentiation is consistent with the role of NOTCH1 as an oncogene in other cancers, especially in T-cell acute lymphoblastic leukemia. We have previously observed that NOTCH1 plays a dual role as both a tumor suppressor and oncogene, depending on the mutational context of the tumor. Namely, gain or loss or NOTCH1 activity promotes the development of human papillomavirus (HPV)-associated cancers. The additional HPV oncogenes likely disrupt the tumor-suppressive activities of NOTCH and enable the oncogenic pathways activated by NOTCH to promote tumor growth. In this review, we detail the role of NOTCH pathway in head and neck cancers with a focus on HPV-associated cancers.
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Structure of High-Risk Papillomavirus 31 E6 Oncogenic Protein and Characterization of E6/E6AP/p53 Complex Formation. J Virol 2020; 95:JVI.00730-20. [PMID: 33115863 DOI: 10.1128/jvi.00730-20] [Citation(s) in RCA: 17] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/21/2020] [Accepted: 10/23/2020] [Indexed: 02/08/2023] Open
Abstract
The degradation of p53 is a hallmark of high-risk human papillomaviruses (HPVs) of the alpha genus and HPV-related carcinogenicity. The oncoprotein E6 forms a ternary complex with the E3 ubiquitin ligase E6-associated protein (E6AP) and tumor suppressor protein p53 targeting p53 for ubiquitination. The extent of p53 degradation by different E6 proteins varies greatly, even for the closely related HPV16 and HPV31. HPV16 E6 and HPV31 E6 display high sequence identity (∼67%). We report here, for the first time, the structure of HPV31 E6 bound to the LxxLL motif of E6AP. HPV16 E6 and HPV31 E6 are structurally very similar, in agreement with the high sequence conservation. Both E6 proteins bind E6AP and degrade p53. However, the binding affinities of 31 E6 to the LxxLL motif of E6AP and p53, respectively, are reduced 2-fold and 5.4-fold compared to 16 E6. The affinity of E6-E6AP-p53 ternary complex formation parallels the efficacy of the subsequent reaction, namely, degradation of p53. Therefore, closely related E6 proteins addressing the same cellular targets may still diverge in their binding efficiencies, possibly explaining their different phenotypic or pathological impacts.IMPORTANCE Variations of carcinogenicity of human papillomaviruses are related to variations of the E6 and E7 interactome. While different HPV species and genera are known to target distinct host proteins, the fine differences between E6 and E7 of closely related HPVs, supposed to target the same cellular protein pools, remain to be addressed. We compare the oncogenic E6 proteins of the closely related high-risk HPV31 and HPV16 with regard to their structure and their efficiency of ternary complex formation with their cellular targets p53 and E6AP, which results in p53 degradation. We solved the crystal structure of 31 E6 bound to the E6AP LxxLL motif. HPV16 E6 and 31 E6 structures are highly similar, but a few sequence variations lead to different protein contacts within the ternary complex and, as quantified here, an overall lower binding affinity of 31 E6 than 16 E6. These results align with the observed lower p53 degradation potential of 31 E6.
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30
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Szymonowicz KA, Chen J. Biological and clinical aspects of HPV-related cancers. Cancer Biol Med 2020; 17:864-878. [PMID: 33299640 PMCID: PMC7721094 DOI: 10.20892/j.issn.2095-3941.2020.0370] [Citation(s) in RCA: 185] [Impact Index Per Article: 37.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/08/2020] [Accepted: 10/21/2020] [Indexed: 12/17/2022] Open
Abstract
Cancer-related diseases represent the second overall cause of death worldwide. Human papilloma virus (HPV) is an infectious agent which is mainly sexually transmitted and may lead to HPV-associated cancers in both men and women. Almost all cervical cancers are HPV-associated, however, an increasing number of head and neck cancers (HNCs), especially oropharyngeal cancer, can be linked to HPV infection. Moreover, anogenital cancers, including vaginal, vulvar, penial, and anal cancers, represent a subset of HPV-related cancers. Whereas testing and prevention of cervical cancer have significantly improved over past decades, anogenital cancers remain more difficult to confirm. Current clinical trials including patients with HPV-related cancers focus on finding proper testing for all HPV-associated cancers as well as improve the currently applied treatments. The HPV viral oncoproteins, E6 and E7, lead to degradation of, respectively, p53 and pRb resulting in entering the S phase without G1 arrest. These high-risk HPV viral oncogenes alter numerous cellular processes, including DNA repair, angiogenesis, and/or apoptosis, which eventually result in carcinogenesis. Additionally, a comprehensive analysis of gene expression and alteration among a panel of DNA double strand breaks (DSB) repair genes in HPV-negative and HPV-positive HNC cancers reveals differences pointing to HPV-dependent modifications of DNA repair processes in these cancers. In this review, we discuss the current knowledge regarding HPV-related cancers, current screening, and treatment options as well as DNA damage response-related biological aspects of the HPV infection and clinical trials.
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Affiliation(s)
- Klaudia Anna Szymonowicz
- Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA
| | - Junjie Chen
- Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA
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31
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The lncRNA H19 alleviates muscular dystrophy by stabilizing dystrophin. Nat Cell Biol 2020; 22:1332-1345. [PMID: 33106653 PMCID: PMC7951180 DOI: 10.1038/s41556-020-00595-5] [Citation(s) in RCA: 54] [Impact Index Per Article: 10.8] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/19/2020] [Accepted: 09/16/2020] [Indexed: 01/04/2023]
Abstract
Dystrophin proteomic regulation in Muscular Dystrophies (MD) remains unclear. We report that a long noncoding RNA (lncRNA), H19, associates with dystrophin and inhibits E3 ligase-dependent poly-ubiquitination at Lys3584 (referred to as Ub-DMD) and its subsequent protein degradation. In-frame deletions in BMD and a DMD non-silent mutation (C3340Y) result in defects in the protein’s ability to interact with H19, causing elevated Ub-DMD levels and dystrophin degradation. Dmd C3333Y mice exhibited progressive muscular dystrophy, elevated serum CK, heart dilation, blood vessel irregularity, and respiratory failure with concurrently reduced dystrophin and increased Ub-DMD status. H19 RNA oligonucleotides conjugated with Agrin (AGR-H19) and Nifenazone competed-with/inhibited TRIM63. Dmd C3333Y animals, iPSC-derived skeletal muscle cells from BMD patients, or mdx mice subjected to exon-skipping exhibited inhibited dystrophin degradation, preserved skeletal/cardiac muscle histology, and improved strength/heart function following AGR-H19 or Nifenazone treatment. Our study paves the way to meaningful targeted therapeutics for BMD and certain DMD patients.
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PKA and Ube3a regulate SK2 channel trafficking to promote synaptic plasticity in hippocampus: Implications for Angelman Syndrome. Sci Rep 2020; 10:9824. [PMID: 32555345 PMCID: PMC7299966 DOI: 10.1038/s41598-020-66790-4] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/13/2019] [Accepted: 05/04/2020] [Indexed: 12/29/2022] Open
Abstract
The ubiquitin ligase, Ube3a, plays important roles in brain development and functions, since its deficiency results in Angelman Syndrome (AS) while its over-expression increases the risk for autism. We previously showed that the lack of Ube3a-mediated ubiquitination of the Ca2+-activated small conductance potassium channel, SK2, contributes to impairment of synaptic plasticity and learning in AS mice. Synaptic SK2 levels are also regulated by protein kinase A (PKA), which phosphorylates SK2 in its C-terminal domain, facilitating its endocytosis. Here, we report that PKA activation restores theta burst stimulation (TBS)-induced long-term potentiation (LTP) in hippocampal slices from AS mice by enhancing SK2 internalization. While TBS-induced SK2 endocytosis is facilitated by PKA activation, SK2 recycling to synaptic membranes after TBS is inhibited by Ube3a. Molecular and cellular studies confirmed that phosphorylation of SK2 in the C-terminal domain increases its ubiquitination and endocytosis. Finally, PKA activation increases SK2 phosphorylation and ubiquitination in Ube3a-overexpressing mice. Our results indicate that, although both Ube3a-mediated ubiquitination and PKA-induced phosphorylation reduce synaptic SK2 levels, phosphorylation is mainly involved in TBS-induced endocytosis, while ubiquitination predominantly inhibits SK2 recycling. Understanding the complex interactions between PKA and Ube3a in the regulation of SK2 synaptic levels might provide new platforms for developing treatments for AS and various forms of autism.
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Karan D, Dubey S, Pirisi L, Nagel A, Pina I, Choo YM, Hamann MT. The Marine Natural Product Manzamine A Inhibits Cervical Cancer by Targeting the SIX1 Protein. JOURNAL OF NATURAL PRODUCTS 2020; 83:286-295. [PMID: 32022559 PMCID: PMC7161578 DOI: 10.1021/acs.jnatprod.9b00577] [Citation(s) in RCA: 36] [Impact Index Per Article: 7.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 07/23/2019] [Indexed: 06/10/2023]
Abstract
Natural products remain an important source of drug leads covering unique chemical space and providing significant therapeutic value for the control of cancer and infectious diseases resistant to current drugs. Here, we determined the antiproliferative activity of a natural product manzamine A (1) from an Indo-Pacific sponge following various in vitro cellular assays targeting cervical cancer (C33A, HeLa, SiHa, and CaSki). Our data demonstrated the antiproliferative effects of 1 at relatively low and non-cytotoxic concentrations (up to 4 μM). Mechanistic investigations confirmed that 1 blocked cell cycle progression in SiHa and CaSki cells at G1/S phase and regulated cell cycle-related genes, including restoration of p21 and p53 expression. In apoptotic assays, HeLa cells showed the highest sensitivity to 1 as compared to other cell types (C33A, SiHa, and CaSki). Interestingly, 1 decreased the levels of the oncoprotein SIX1, which is associated with oncogenesis in cervical cancer. To further investigate the structure-activity relationship among manzamine A (1) class with potential antiproliferative activity, molecular networking facilitated the efficient identification, dereplication, and assignment of structures from the manzamine class and revealed the significant potential in the design of optimized molecules for the treatment of cervical cancer. These data suggest that this sponge-derived natural product class warrants further attention regarding the design and development of novel manzamine analogues, which may be efficacious for preventive and therapeutic treatment of cancer. Additionally, this study reveals the significance of protecting fragile marine ecosystems from climate change-induced loss of species diversity.
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Affiliation(s)
- Dev Karan
- Department
of Pathology, MCW Cancer Center and Prostate Cancer Center of Excellence, Medical College of Wisconsin, 8701 Watertown Plank Road, Milwaukee, Wisconsin, United States
| | - Seema Dubey
- Department
of Pathology, MCW Cancer Center and Prostate Cancer Center of Excellence, Medical College of Wisconsin, 8701 Watertown Plank Road, Milwaukee, Wisconsin, United States
| | - Lucia Pirisi
- Department
of Pathology, Microbiology and Immunology, University of South Carolina School of Medicine, Columbia, South Carolina, United States
| | - Alexis Nagel
- Department
of Drug Discovery and Biomedical Sciences, Medical University of South Carolina, Charleston, South Carolina, United States
| | - Ivett Pina
- Department
of Drug Discovery and Biomedical Sciences, Medical University of South Carolina, Charleston, South Carolina, United States
| | - Yeun-Mun Choo
- Department
of Chemistry, University of Malaya, Kuala Lumpur, Malaysia
| | - Mark T Hamann
- Department
of Drug Discovery and Biomedical Sciences, Medical University of South Carolina, Charleston, South Carolina, United States
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Rotaru DC, Mientjes EJ, Elgersma Y. Angelman Syndrome: From Mouse Models to Therapy. Neuroscience 2020; 445:172-189. [PMID: 32088294 DOI: 10.1016/j.neuroscience.2020.02.017] [Citation(s) in RCA: 42] [Impact Index Per Article: 8.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2019] [Revised: 02/10/2020] [Accepted: 02/11/2020] [Indexed: 12/19/2022]
Abstract
The UBE3A gene is part of the chromosome 15q11-q13 region that is frequently deleted or duplicated, leading to several neurodevelopmental disorders (NDD). Angelman syndrome (AS) is caused by the absence of functional maternally derived UBE3A protein, while the paternal UBE3A gene is present but silenced specifically in neurons. Patients with AS present with severe neurodevelopmental delay, with pronounced motor deficits, absence of speech, intellectual disability, epilepsy, and sleep problems. The pathophysiology of AS is still unclear and a treatment is lacking. Animal models of AS recapitulate the genotypic and phenotypic features observed in AS patients, and have been invaluable for understanding the disease process as well as identifying apropriate drug targets. Using these AS mouse models we have learned that loss of UBE3A probably affects many areas of the brain, leading to increased neuronal excitability and a loss of synaptic spines, along with changes in a number of distinct behaviours. Inducible AS mouse models have helped to identify the critical treatment windows for the behavioral and physiological phenotypes. Additionally, AS mouse models indicate an important role for the predominantly nuclear UBE3A isoform in generating the characteristic AS pathology. Last, but not least, the AS mice have been crucial in guiding Ube3a gene reactivation treatments, which present a very promising therapy to treat AS.
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Affiliation(s)
- Diana C Rotaru
- Department of Neuroscience, The ENCORE Expertise Center for Neurodevelopmental Disorders, Erasmus MC University Medical Center, Rotterdam, The Netherlands
| | - Edwin J Mientjes
- Department of Neuroscience, The ENCORE Expertise Center for Neurodevelopmental Disorders, Erasmus MC University Medical Center, Rotterdam, The Netherlands
| | - Ype Elgersma
- Department of Neuroscience, The ENCORE Expertise Center for Neurodevelopmental Disorders, Erasmus MC University Medical Center, Rotterdam, The Netherlands.
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Đukić A, Lulić L, Thomas M, Skelin J, Bennett Saidu NE, Grce M, Banks L, Tomaić V. HPV Oncoproteins and the Ubiquitin Proteasome System: A Signature of Malignancy? Pathogens 2020; 9:pathogens9020133. [PMID: 32085533 PMCID: PMC7168213 DOI: 10.3390/pathogens9020133] [Citation(s) in RCA: 25] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/20/2019] [Revised: 02/10/2020] [Accepted: 02/11/2020] [Indexed: 12/22/2022] Open
Abstract
Human papillomavirus (HPV) E6 and E7 oncoproteins are critical for development and maintenance of the malignant phenotype in HPV-induced cancers. These two viral oncoproteins interfere with a plethora of cellular pathways, including the regulation of cell cycle and the control of apoptosis, which are critical in maintaining normal cellular functions. E6 and E7 bind directly with certain components of the Ubiquitin Proteasome System (UPS), enabling them to manipulate a number of important cellular pathways. These activities are the means by which HPV establishes an environment supporting the normal viral life cycle, however in some instances they can also lead to the development of malignancy. In this review, we have discussed how E6 and E7 oncoproteins from alpha and beta HPV types interact with the components of the UPS, and how this interplay contributes to the development of cancer.
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Affiliation(s)
- Anamaria Đukić
- Division of Molecular Medicine, Ruđer Bošković Institute, Bijenička cesta 54, 10000 Zagreb, Croatia; (A.Đ.); (L.L.); (J.S.); (N.E.B.S.); (M.G.)
| | - Lucija Lulić
- Division of Molecular Medicine, Ruđer Bošković Institute, Bijenička cesta 54, 10000 Zagreb, Croatia; (A.Đ.); (L.L.); (J.S.); (N.E.B.S.); (M.G.)
| | - Miranda Thomas
- International Centre for Genetic Engineering and Biotechnology, AREA Science Park, Padriciano 99, I-34149 Trieste, Italy; (M.T.); (L.B.)
| | - Josipa Skelin
- Division of Molecular Medicine, Ruđer Bošković Institute, Bijenička cesta 54, 10000 Zagreb, Croatia; (A.Đ.); (L.L.); (J.S.); (N.E.B.S.); (M.G.)
| | - Nathaniel Edward Bennett Saidu
- Division of Molecular Medicine, Ruđer Bošković Institute, Bijenička cesta 54, 10000 Zagreb, Croatia; (A.Đ.); (L.L.); (J.S.); (N.E.B.S.); (M.G.)
| | - Magdalena Grce
- Division of Molecular Medicine, Ruđer Bošković Institute, Bijenička cesta 54, 10000 Zagreb, Croatia; (A.Đ.); (L.L.); (J.S.); (N.E.B.S.); (M.G.)
| | - Lawrence Banks
- International Centre for Genetic Engineering and Biotechnology, AREA Science Park, Padriciano 99, I-34149 Trieste, Italy; (M.T.); (L.B.)
| | - Vjekoslav Tomaić
- Division of Molecular Medicine, Ruđer Bošković Institute, Bijenička cesta 54, 10000 Zagreb, Croatia; (A.Đ.); (L.L.); (J.S.); (N.E.B.S.); (M.G.)
- Correspondence: ; Tel.: +385-1-4561110; Fax: +385-1-4561010
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Lo Cigno I, Calati F, Borgogna C, Zevini A, Albertini S, Martuscelli L, De Andrea M, Hiscott J, Landolfo S, Gariglio M. Human Papillomavirus E7 Oncoprotein Subverts Host Innate Immunity via SUV39H1-Mediated Epigenetic Silencing of Immune Sensor Genes. J Virol 2020; 94:e01812-19. [PMID: 31776268 PMCID: PMC6997746 DOI: 10.1128/jvi.01812-19] [Citation(s) in RCA: 45] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/28/2019] [Accepted: 11/20/2019] [Indexed: 12/23/2022] Open
Abstract
Subversion of innate immunity by oncoviruses, such as human papillomavirus (HPV), favors carcinogenesis because the mechanism(s) of viral immune evasion can also hamper cancer immunosurveillance. Previously, we demonstrated that high-risk (hr) HPVs trigger simultaneous epigenetic silencing of multiple effectors of innate immunity to promote viral persistence. Here, we expand on those observations and show that the HPV E7 oncoprotein upregulates the H3K9-specific methyltransferase, whose action shuts down the host innate immune response. Specifically, we demonstrate that SUV39H1 contributes to chromatin repression at the promoter regions of the viral nucleic acid sensors RIG-I and cGAS and the adaptor molecule STING in HPV-transformed cells. Inhibition of SUV39H1 leads to transcriptional activation of these genes, especially RIG-I, followed by increased beta interferon (IFN-β) and IFN-λ1 production after poly(dA·dT) or RIG-I agonist M8 transfection. Collectively, our findings provide new evidence that the E7 oncoprotein plays a central role in dampening host innate immunity and raise the possibility that targeting the downstream effector SUV39H1 or the RIG-I pathway is a viable strategy to treat viral and neoplastic disease.IMPORTANCE High-risk HPVs are major viral human carcinogens responsible for approximately 5% of all human cancers. The growth of HPV-transformed cells depends on the ability of viral oncoproteins to manipulate a variety of cellular circuits, including those involved in innate immunity. Here, we show that one of these strategies relies on E7-mediated transcriptional activation of the chromatin repressor SUV39H1, which then promotes epigenetic silencing of RIG-I, cGAS, and STING genes, thereby shutting down interferon secretion in HPV-transformed cells. Pharmacological or genetic inhibition of SUV39H1 restored the innate response in HPV-transformed cells, mostly through activation of RIG-I signaling. We also show that IFN production upon transfection of poly(dA·dT) or the RIG-I agonist M8 predominantly occurs through RIG-I signaling. Altogether, the reversible nature of the modifications associated with E7-mediated SUV39H1 upregulation provides a rationale for the design of novel anticancer and antiviral therapies targeting these molecules.
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Affiliation(s)
- Irene Lo Cigno
- University of Piemonte Orientale Medical School, Department of Translational Medicine, Molecular Virology Unit, Novara, Italy
| | - Federica Calati
- University of Piemonte Orientale Medical School, Department of Translational Medicine, Molecular Virology Unit, Novara, Italy
| | - Cinzia Borgogna
- University of Piemonte Orientale Medical School, Department of Translational Medicine, Molecular Virology Unit, Novara, Italy
| | | | - Silvia Albertini
- University of Piemonte Orientale Medical School, Department of Translational Medicine, Molecular Virology Unit, Novara, Italy
| | - Licia Martuscelli
- University of Piemonte Orientale Medical School, Department of Translational Medicine, Molecular Virology Unit, Novara, Italy
| | - Marco De Andrea
- Center for Translational Research on Autoimmune and Allergic Disease-CAAD, Novara, Italy
- University of Turin Medical School, Department of Public Health and Pediatric Sciences, Viral Pathogenesis Unit, Turin, Italy
| | - John Hiscott
- Istituto Pasteur-Fondazione Cenci Bolognetti, Rome, Italy
| | - Santo Landolfo
- University of Turin Medical School, Department of Public Health and Pediatric Sciences, Viral Pathogenesis Unit, Turin, Italy
| | - Marisa Gariglio
- University of Piemonte Orientale Medical School, Department of Translational Medicine, Molecular Virology Unit, Novara, Italy
- Center for Translational Research on Autoimmune and Allergic Disease-CAAD, Novara, Italy
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37
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Pyeon D, Rojas VK, Price L, Kim S, Singh M, Park IW. HIV-1 Impairment via UBE3A and HIV-1 Nef Interactions Utilizing the Ubiquitin Proteasome System. Viruses 2019; 11:v11121098. [PMID: 31783587 PMCID: PMC6950590 DOI: 10.3390/v11121098] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/21/2019] [Revised: 11/15/2019] [Accepted: 11/25/2019] [Indexed: 12/30/2022] Open
Abstract
Molecular basis of HIV-1 life cycle regulation has thus far focused on viral gene stage-specificity, despite the quintessence of post-function protein elimination processes in the virus life cycle and consequent pathogenesis. Our studies demonstrated that a key pathogenic HIV-1 viral protein, Nef, interacted with ubiquitin (Ub)-protein ligase E3A (UBE3A/E6AP), suggesting that interaction between Nef and UBE3A is integral to regulation of viral and cellular protein decay and thereby the competing HIV-1 and host cell survivals. In fact, Nef and UBE3A degraded reciprocally, and UBE3A-mediated degradation of Nef was significantly more potent than Nef-triggered degradation of UBE3A. Further, UBE3A degraded not only Nef but also HIV-1 structural proteins, Gag, thus significantly inhibiting HIV-1 replication in Jurkat T cells only in the presence of Nef, indicating that interaction between Nef and UBE3Awas pivotal for UBE3A-mediated degradation of the viral proteins. Mechanistic study showed that Nef and UBE3A were specific and antagonistic to each other in regulating proteasome activity and ubiquitination of cellular proteins in general, wherein specific domains of Nef overlapping with the long terminal repeat (LTR) were essential for the observed actions. Further, Nef itself reduced the level of intracellular Gag by degrading a cardinal transcription regulator, Tat, demonstrating a broad role for Nef in the regulation of the HIV-1 life cycle. Taken together, these data demonstrated that the Nef and UBE3A complex plays a crucial role in coordinating viral protein degradation and hence HIV-1 replication, providing insights as to the nature of pathobiologic and defense strategies of HIV-1 and HIV-infected host cells.
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Affiliation(s)
- Dohun Pyeon
- Departments of Microbiology and Molecular Genetics, Michigan State University, East Lansing, MI 48824, USA;
| | - Vivian K. Rojas
- Department of Microbiology, Immunology and Genetics, University of North Texas Health Science Center, Fort Worth, TX 76107, USA; (V.K.R.); (L.P.)
| | - Lenore Price
- Department of Microbiology, Immunology and Genetics, University of North Texas Health Science Center, Fort Worth, TX 76107, USA; (V.K.R.); (L.P.)
| | - Seongcheol Kim
- Pharmacology and Neuroscience, University of North Texas Health Science Center, Fort Worth, TX 76107, USA (M.S.)
| | - Meharvan Singh
- Pharmacology and Neuroscience, University of North Texas Health Science Center, Fort Worth, TX 76107, USA (M.S.)
| | - In-Woo Park
- Department of Microbiology, Immunology and Genetics, University of North Texas Health Science Center, Fort Worth, TX 76107, USA; (V.K.R.); (L.P.)
- Correspondence: ; Tel.: +(817)-735-5115; Fax: +(817)-735-2610
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Vats A, Thatte J, Banks L. Identification of E6AP-independent degradation targets of HPV E6. J Gen Virol 2019; 100:1674-1679. [PMID: 31609195 DOI: 10.1099/jgv.0.001331] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
The high-risk Human Papillomavirus (HPV) E6 oncoprotein is known to contribute to human malignancy by targeting several of its cellular substrates through the ubiquitin-mediated degradation pathway. Previous studies have revealed that E6 interacts with the E6AP ubiquitin-protein ligase and directs its ubiquitylation activity toward several specific cellular proteins, one of the most important of which is p53. However, the role of E6AP in the degradation of many other E6 substrates is still ambiguous because loss of E6AP also induces a loss of E6 expression. To examine this further, we used CRISPR-edited E6AP knockout cells to perform E6 degradation assays in the presence of a catalytically inactive mutant form of E6AP, thus ensuring the stabilization of E6 but with the ligase itself being functionally inactive. Using this system, we found that E6 can mediate the degradation of several PDZ domain-containing proteins independently of E6AP ubiquitin ligase activity. This study thus opens up ways to investigate other possible components of the cellular ubiquitin proteasome pathway that E6 might utilize to target these substrates.
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Affiliation(s)
- Arushi Vats
- International Centre for Genetic Engineering and Biotechnology, Padriciano 99 I-34149 Trieste, Italy
| | - Jayashree Thatte
- International Centre for Genetic Engineering and Biotechnology, Padriciano 99 I-34149 Trieste, Italy
| | - Lawrence Banks
- International Centre for Genetic Engineering and Biotechnology, Padriciano 99 I-34149 Trieste, Italy
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39
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Mishra M, Sharma A, Thacker G, Trivedi AK. Nano-LC based proteomic approach identifies that E6AP interacts with ENO1 and targets it for degradation in breast cancer cells. IUBMB Life 2019; 71:1896-1905. [PMID: 31329371 DOI: 10.1002/iub.2132] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/24/2019] [Accepted: 07/01/2019] [Indexed: 12/11/2022]
Abstract
E6AP (E6 associated protein) is a HECT domain containing protein having dual E3 ligase and ERα coactivation activity in breast cancer cells. Although E6AP is known to possess antitumorigenic activity, the underlying molecular mechanism is poorly understood. In the present study, we applied nano-LC based proteomics approach to identify E6AP-interacting proteins where we performed GST-pull down using GST-E6AP from whole cell extracts of MCF7 cells, resolved the differentially interacting proteins on 1D-SDS-PAGE, excised the gel bands that were trypsin digested followed by fractionation and spotting on MALDI-TOF/TOF plate through Nano-LC MALDI spotter. Subsequently, fractionated and spotted peptides were identified using MALDI-TOF/TOF. We identified several E6AP interacting proteins including previously reported such as HSP70 and new ones such as Enolase-1. We further confirmed that E6AP and Enolase1 interacted and colocalized more in the cytoplasmic periphery in breast cancer cells and further demonstrated that E6AP also targeted ENO1 for ubiquitin-mediated degradation in these cells.
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Affiliation(s)
- Mukul Mishra
- Room No. LSS008, Biochemistry Division, CSIR-Central Drug Research Institute, Lucknow, Uttar Pradesh, India
| | - Akshay Sharma
- Room No. LSS008, Biochemistry Division, CSIR-Central Drug Research Institute, Lucknow, Uttar Pradesh, India
| | - Gatha Thacker
- Room No. LSS008, Biochemistry Division, CSIR-Central Drug Research Institute, Lucknow, Uttar Pradesh, India
| | - Arun K Trivedi
- Room No. LSS008, Biochemistry Division, CSIR-Central Drug Research Institute, Lucknow, Uttar Pradesh, India.,Academy of Scientific and Innovative Research (AcSIR), CSIR-Central Drug Research Institute, Lucknow, Uttar Pradesh, India
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40
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Kim J, Lee B, Kim DH, Yeon JG, Lee J, Park Y, Lee Y, Lee SK, Lee S, Lee JW. UBE3A Suppresses Overnutrition-Induced Expression of the Steatosis Target Genes of MLL4 by Degrading MLL4. Hepatology 2019; 69:1122-1134. [PMID: 30230575 PMCID: PMC6393921 DOI: 10.1002/hep.30284] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/22/2018] [Accepted: 09/05/2018] [Indexed: 12/28/2022]
Abstract
Regulation of the protein stability of epigenetic regulators remains ill-defined despite its potential applicability in epigenetic therapies. The histone H3-lysine 4-methyltransferase MLL4 is an epigenetic transcriptional coactivator that directs overnutrition-induced obesity and fatty liver formation, and Mll4+/- mice are resistant to both. Here we show that the E3 ubiquitin ligase UBE3A targets MLL4 for degradation, thereby suppressing high-fat diet (HFD)-induced expression of the hepatic steatosis target genes of MLL4. In contrast to Mll4+/- mice, Ube3a+/- mice are hypersensitive to HFD-induced obesity and fatty liver development. Ube3a+/-;Mll4+/- mice lose this hypersensitivity, supporting roles of increased MLL4 levels in both phenotypes of Ube3a+/- mice. Correspondingly, our comparative studies with wild-type, Ube3a+/- and Ube3a-/- and UBE3A-overexpressing transgenic mouse livers demonstrate an inverse correlation of UBE3A protein levels with MLL4 protein levels, expression of the steatosis target genes of MLL4, and their decoration by H3-lysine 4-monomethylation, a surrogate marker for the epigenetic action of MLL4. Conclusion: UBE3A indirectly exerts an epigenetic regulation of obesity and steatosis by degrading MLL4. This UBE3A-MLL4 regulatory axis provides a potential therapeutic venue for treating various MLL4-directed pathogeneses, including obesity and hepatic steatosis.
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Affiliation(s)
- Janghyun Kim
- Neuroscience Section, Papé Family Pediatric Research
Institute, Department of Pediatrics, Oregon Health & Science University,
Portland, OR 97239, USA
| | - Bora Lee
- Center for Neuroscience, Korea Institute of Science and
Technology, Seoul 02792, Korea
| | - Dae-Hwan Kim
- Neuroscience Section, Papé Family Pediatric Research
Institute, Department of Pediatrics, Oregon Health & Science University,
Portland, OR 97239, USA
| | - Je Gwang Yeon
- College of Pharmacy and Research Institute of
Pharmaceutical Sciences, Seoul National University, Seoul 08826, Korea
| | - Jeongkyung Lee
- Division of Endocrinology & Metabolism, Department of
Medicine, University of Pittsburgh, Pittsburgh, PA 15213, USA
| | - Younjung Park
- Neuroscience Section, Papé Family Pediatric Research
Institute, Department of Pediatrics, Oregon Health & Science University,
Portland, OR 97239, USA
| | - Yuna Lee
- Neuroscience Section, Papé Family Pediatric Research
Institute, Department of Pediatrics, Oregon Health & Science University,
Portland, OR 97239, USA
| | - Soo-Kyung Lee
- Neuroscience Section, Papé Family Pediatric Research
Institute, Department of Pediatrics, Oregon Health & Science University,
Portland, OR 97239, USA,Vollum Institute, Oregon Health & Science University,
Portland, OR 97239, USA
| | - Seunghee Lee
- College of Pharmacy and Research Institute of
Pharmaceutical Sciences, Seoul National University, Seoul 08826, Korea,Correspondences: Seunghee Lee
() or Jae W. Lee
()
| | - Jae W. Lee
- Neuroscience Section, Papé Family Pediatric Research
Institute, Department of Pediatrics, Oregon Health & Science University,
Portland, OR 97239, USA,Correspondences: Seunghee Lee
() or Jae W. Lee
()
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41
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Rodriguez J, Herrero A, Li S, Rauch N, Quintanilla A, Wynne K, Krstic A, Acosta JC, Taylor C, Schlisio S, von Kriegsheim A. PHD3 Regulates p53 Protein Stability by Hydroxylating Proline 359. Cell Rep 2018; 24:1316-1329. [PMID: 30067985 PMCID: PMC6088137 DOI: 10.1016/j.celrep.2018.06.108] [Citation(s) in RCA: 53] [Impact Index Per Article: 7.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/04/2017] [Revised: 03/30/2018] [Accepted: 06/27/2018] [Indexed: 01/05/2023] Open
Abstract
Cellular p53 protein levels are regulated by a ubiquitination/de-ubiquitination cycle that can target the protein for proteasomal destruction. The ubiquitination reaction is catalyzed by a multitude of ligases, whereas the removal of ubiquitin chains is mediated by two deubiquitinating enzymes (DUBs), USP7 (HAUSP) and USP10. Here, we show that PHD3 hydroxylates p53 at proline 359, a residue that is in the p53-DUB binding domain. Hydroxylation of p53 upon proline 359 regulates its interaction with USP7 and USP10, and its inhibition decreases the association of p53 with USP7/USP10, increases p53 ubiquitination, and rapidly reduces p53 protein levels independently of mRNA expression. Our results show that p53 is a PHD3 substrate and that hydroxylation by PHD3 regulates p53 protein stability through modulation of ubiquitination.
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Affiliation(s)
- Javier Rodriguez
- Systems Biology Ireland, University College Dublin, Dublin 4, Ireland; Cancer Research UK Edinburgh Centre, IGMM, University of Edinburgh, Edinburgh EH4 2XR, UK
| | - Ana Herrero
- Systems Biology Ireland, University College Dublin, Dublin 4, Ireland
| | - Shuijie Li
- Ludwig Institute for Cancer Research Ltd., SE-17177 Stockholm, Sweden; Department of Microbiology and Tumor and Cell Biology, Karolinska Institutet, SE-17177 Stockholm, Sweden
| | - Nora Rauch
- Systems Biology Ireland, University College Dublin, Dublin 4, Ireland
| | - Andrea Quintanilla
- Cancer Research UK Edinburgh Centre, IGMM, University of Edinburgh, Edinburgh EH4 2XR, UK
| | - Kieran Wynne
- Conway Institute, University College Dublin, Dublin 4, Ireland
| | - Aleksandar Krstic
- Systems Biology Ireland, University College Dublin, Dublin 4, Ireland
| | - Juan Carlos Acosta
- Cancer Research UK Edinburgh Centre, IGMM, University of Edinburgh, Edinburgh EH4 2XR, UK
| | - Cormac Taylor
- Conway Institute, University College Dublin, Dublin 4, Ireland
| | - Susanne Schlisio
- Ludwig Institute for Cancer Research Ltd., SE-17177 Stockholm, Sweden; Department of Microbiology and Tumor and Cell Biology, Karolinska Institutet, SE-17177 Stockholm, Sweden
| | - Alex von Kriegsheim
- Systems Biology Ireland, University College Dublin, Dublin 4, Ireland; Cancer Research UK Edinburgh Centre, IGMM, University of Edinburgh, Edinburgh EH4 2XR, UK.
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42
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Sun J, Liu Y, Jia Y, Hao X, Lin WJ, Tran J, Lynch G, Baudry M, Bi X. UBE3A-mediated p18/LAMTOR1 ubiquitination and degradation regulate mTORC1 activity and synaptic plasticity. eLife 2018; 7:37993. [PMID: 30020076 PMCID: PMC6063731 DOI: 10.7554/elife.37993] [Citation(s) in RCA: 32] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/30/2018] [Accepted: 07/17/2018] [Indexed: 01/04/2023] Open
Abstract
Accumulating evidence indicates that the lysosomal Ragulator complex is essential for full activation of the mechanistic target of rapamycin complex 1 (mTORC1). Abnormal mTORC1 activation has been implicated in several developmental neurological disorders, including Angelman syndrome (AS), which is caused by maternal deficiency of the ubiquitin E3 ligase UBE3A. Here we report that Ube3a regulates mTORC1 signaling by targeting p18, a subunit of the Ragulator. Ube3a ubiquinates p18, resulting in its proteasomal degradation, and Ube3a deficiency in the hippocampus of AS mice induces increased lysosomal localization of p18 and other members of the Ragulator-Rag complex, and increased mTORC1 activity. p18 knockdown in hippocampal CA1 neurons of AS mice reduces elevated mTORC1 activity and improves dendritic spine maturation, long-term potentiation (LTP), as well as learning performance. Our results indicate that Ube3a-mediated regulation of p18 and subsequent mTORC1 signaling is critical for typical synaptic plasticity, dendritic spine development, and learning and memory.
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Affiliation(s)
- Jiandong Sun
- Department of Basic Medical Sciences, College of Osteopathic Medicine of the Pacific, Western University of Health Sciences, Pomona, United States
| | - Yan Liu
- Graduate College of Biomedical Sciences, Western University of Health Sciences, Pomona, United States
| | - Yousheng Jia
- Department of Psychiatry, University of California, Irvine, United States
| | - Xiaoning Hao
- Department of Basic Medical Sciences, College of Osteopathic Medicine of the Pacific, Western University of Health Sciences, Pomona, United States
| | - Wei Ju Lin
- Department of Basic Medical Sciences, College of Osteopathic Medicine of the Pacific, Western University of Health Sciences, Pomona, United States
| | - Jennifer Tran
- Department of Basic Medical Sciences, College of Osteopathic Medicine of the Pacific, Western University of Health Sciences, Pomona, United States
| | - Gary Lynch
- Department of Psychiatry, University of California, Irvine, United States
| | - Michel Baudry
- Graduate College of Biomedical Sciences, Western University of Health Sciences, Pomona, United States
| | - Xiaoning Bi
- Department of Basic Medical Sciences, College of Osteopathic Medicine of the Pacific, Western University of Health Sciences, Pomona, United States
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Raghu D, Paul PJ, Gulati T, Deb S, Khoo C, Russo A, Gallo E, Blandino G, Chan AL, Takano E, Sandhu SK, Fox SB, Williams S, Haupt S, Gamell C, Haupt Y. E6AP promotes prostate cancer by reducing p27 expression. Oncotarget 2018; 8:42939-42948. [PMID: 28477016 PMCID: PMC5522117 DOI: 10.18632/oncotarget.17224] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2017] [Accepted: 03/16/2017] [Indexed: 12/03/2022] Open
Abstract
Prostate cancer (PC) is the most common cancer in men. Elevated levels of E3 ligase, E6-Associated Protein (E6AP) were previously linked to PC, consistent with increased protein expression in a subset of PC patients. In cancers, irregular E3 ligase activity drives proteasomal degradation of tumor suppressor proteins. Accordingly, E3 ligase inhibitors define a rational therapy to restore tumor suppression. The relevant tumor suppressors targeted by E6AP in PC are yet to be fully identified. In this study we show that p27, a key cell cycle regulator, is a target of E6AP in PC. Down regulation of E6AP increases p27 expression and enhances its nuclear accumulation in PC. We demonstrate that E6AP regulates p27 expression by inhibiting its transcription in an E2F1-dependent manner. Concomitant knockdown of E6AP and p27 partially restores PC cell growth, supporting the contribution of p27 to the overall effect of E6AP on prostate tumorigenesis. Overall, we unravelled the E6AP-p27 axis as a new promoter of PC, exposing an attractive target for therapy through the restoration of tumor suppression.
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Affiliation(s)
- Dinesh Raghu
- The Sir Peter MacCallum Department of Oncology, The University of Melbourne, Melbourne, Victoria, Australia.,Tumor Suppression Laboratory, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia
| | - Piotr Jan Paul
- The Sir Peter MacCallum Department of Oncology, The University of Melbourne, Melbourne, Victoria, Australia.,Tumor Suppression Laboratory, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia
| | - Twishi Gulati
- The Sir Peter MacCallum Department of Oncology, The University of Melbourne, Melbourne, Victoria, Australia.,Tumor Suppression Laboratory, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia
| | - Siddhartha Deb
- Anatpath Services Pty Ltd, Gardenvale, Victoria, Australia
| | - Christine Khoo
- Department of Pathology, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia
| | - Andrea Russo
- Department of Surgical Pathology, Regina Elena Cancer Institute, Rome, Italy
| | - Enzo Gallo
- Department of Surgical Pathology, Regina Elena Cancer Institute, Rome, Italy
| | - Giovanni Blandino
- Oncogenomic and Epigenetic Unit, Italian National Cancer Institute, Rome, Italy
| | - Ai-Leen Chan
- Tumor Suppression Laboratory, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.,Current address: Australian Regenerative Medicine Institute, Monash University, Melbourne, Victoria, Australia
| | - Elena Takano
- Department of Pathology, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia
| | - Shahneen K Sandhu
- Division of Cancer Medicine, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia
| | - Stephen B Fox
- Department of Pathology, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia
| | - Scott Williams
- Division of Radiation Oncology and Cancer Imaging, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia
| | - Sue Haupt
- The Sir Peter MacCallum Department of Oncology, The University of Melbourne, Melbourne, Victoria, Australia.,Tumor Suppression Laboratory, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia
| | - Cristina Gamell
- The Sir Peter MacCallum Department of Oncology, The University of Melbourne, Melbourne, Victoria, Australia.,Tumor Suppression Laboratory, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia
| | - Ygal Haupt
- The Sir Peter MacCallum Department of Oncology, The University of Melbourne, Melbourne, Victoria, Australia.,Tumor Suppression Laboratory, Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia.,Department of Biochemistry and Molecular Biology, Monash University, Melbourne, Victoria, Australia.,Department of Pathology, The University of Melbourne, Melbourne, Victoria, Australia
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44
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Romeo M, Hutchison T, Malu A, White A, Kim J, Gardner R, Smith K, Nelson K, Bergeson R, McKee R, Harrod C, Ratner L, Lüscher B, Martinez E, Harrod R. The human T-cell leukemia virus type-1 p30 II protein activates p53 and induces the TIGAR and suppresses oncogene-induced oxidative stress during viral carcinogenesis. Virology 2018; 518:103-115. [PMID: 29462755 DOI: 10.1016/j.virol.2018.02.010] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/08/2017] [Revised: 01/23/2018] [Accepted: 02/09/2018] [Indexed: 01/08/2023]
Abstract
In normal cells, aberrant oncogene expression leads to the accumulation of cytotoxic metabolites, including reactive oxygen species (ROS), which can cause oxidative DNA-damage and apoptosis as an intrinsic barrier against neoplastic disease. The c-Myc oncoprotein is overexpressed in many lymphoid cancers due to c-myc gene amplification and/or 8q24 chromosomal translocations. Intriguingly, p53 is a downstream target of c-Myc and hematological malignancies, such as adult T-cell leukemia/lymphoma (ATL), frequently contain wildtype p53 and c-Myc overexpression. We therefore hypothesized that p53-regulated pro-survival signals may thwart the cell's metabolic anticancer defenses to support oncogene-activation in lymphoid cancers. Here we show that the Tp53-induced glycolysis and apoptosis regulator (TIGAR) promotes c-myc oncogene-activation by the human T-cell leukemia virus type-1 (HTLV-1) latency-maintenance factor p30II, associated with c-Myc deregulation in ATL clinical isolates. TIGAR prevents the intracellular accumulation of c-Myc-induced ROS and inhibits oncogene-induced cellular senescence in ATL, acute lymphoblastic leukemia, and multiple myeloma cells with elevated c-Myc expression. Our results allude to a pivotal role for p53-regulated antioxidant signals as mediators of c-Myc oncogenic functions in viral and non-viral lymphoid tumors.
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Affiliation(s)
- Megan Romeo
- Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design & Delivery, Southern Methodist University, Dallas, TX 75275-0376, Unites States
| | - Tetiana Hutchison
- Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design & Delivery, Southern Methodist University, Dallas, TX 75275-0376, Unites States
| | - Aditi Malu
- Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design & Delivery, Southern Methodist University, Dallas, TX 75275-0376, Unites States
| | - Averi White
- Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design & Delivery, Southern Methodist University, Dallas, TX 75275-0376, Unites States
| | - Janice Kim
- Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design & Delivery, Southern Methodist University, Dallas, TX 75275-0376, Unites States
| | - Rachel Gardner
- Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design & Delivery, Southern Methodist University, Dallas, TX 75275-0376, Unites States
| | - Katie Smith
- Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design & Delivery, Southern Methodist University, Dallas, TX 75275-0376, Unites States
| | - Katherine Nelson
- Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design & Delivery, Southern Methodist University, Dallas, TX 75275-0376, Unites States
| | - Rachel Bergeson
- Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design & Delivery, Southern Methodist University, Dallas, TX 75275-0376, Unites States
| | - Ryan McKee
- Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design & Delivery, Southern Methodist University, Dallas, TX 75275-0376, Unites States
| | - Carolyn Harrod
- Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design & Delivery, Southern Methodist University, Dallas, TX 75275-0376, Unites States
| | - Lee Ratner
- Departments of Medicine and Molecular Microbiology, Washington University School of Medicine, St. Louis, MO 63110, United States
| | - Bernhard Lüscher
- Institute of Biochemistry, Klinikum, RWTH Aachen University, Pauwelsstrasse 30, 52057 Aachen, Germany
| | - Ernest Martinez
- Department of Biochemistry, University of California, Riverside, CA 92521, United States
| | - Robert Harrod
- Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design & Delivery, Southern Methodist University, Dallas, TX 75275-0376, Unites States.
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Olmedo-Nieva L, Muñoz-Bello JO, Contreras-Paredes A, Lizano M. The Role of E6 Spliced Isoforms (E6*) in Human Papillomavirus-Induced Carcinogenesis. Viruses 2018; 10:v10010045. [PMID: 29346309 PMCID: PMC5795458 DOI: 10.3390/v10010045] [Citation(s) in RCA: 50] [Impact Index Per Article: 7.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/22/2017] [Revised: 01/12/2018] [Accepted: 01/15/2018] [Indexed: 12/13/2022] Open
Abstract
Persistent infections with High Risk Human Papillomaviruses (HR-HPVs) are the main cause of cervical cancer development. The E6 and E7 oncoproteins of HR-HPVs are derived from a polycistronic pre-mRNA transcribed from an HPV early promoter. Through alternative splicing, this pre-mRNA produces a variety of E6 spliced transcripts termed E6*. In pre-malignant lesions and HPV-related cancers, different E6/E6* transcriptional patterns have been found, although they have not been clearly associated to cancer development. Moreover, there is a controversy about the participation of E6* proteins in cancer progression. This review addresses the regulation of E6 splicing and the different functions that have been found for E6* proteins, as well as their possible role in HPV-induced carcinogenesis.
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Affiliation(s)
- Leslie Olmedo-Nieva
- Unidad de Investigación Biomédica en Cáncer, Instituto Nacional de Cancerología, México/Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Av. San Fernando No. 22, Col. Sección XVI, Tlalpan, 14080 Mexico City, Mexico.
| | - J Omar Muñoz-Bello
- Unidad de Investigación Biomédica en Cáncer, Instituto Nacional de Cancerología, México/Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Av. San Fernando No. 22, Col. Sección XVI, Tlalpan, 14080 Mexico City, Mexico.
| | - Adriana Contreras-Paredes
- Unidad de Investigación Biomédica en Cáncer, Instituto Nacional de Cancerología, México/Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Av. San Fernando No. 22, Col. Sección XVI, Tlalpan, 14080 Mexico City, Mexico.
| | - Marcela Lizano
- Unidad de Investigación Biomédica en Cáncer, Instituto Nacional de Cancerología, México/Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Av. San Fernando No. 22, Col. Sección XVI, Tlalpan, 14080 Mexico City, Mexico.
- Departamento de Medicina Genómica y Toxicología Ambiental, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, 04510 Mexico City, Mexico.
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DasGupta T, Nweze EI, Yue H, Wang L, Jin J, Ghosh SK, Kawsar HI, Zender C, Androphy EJ, Weinberg A, McCormick TS, Jin G. Human papillomavirus oncogenic E6 protein regulates human β-defensin 3 (hBD3) expression via the tumor suppressor protein p53. Oncotarget 2017; 7:27430-44. [PMID: 27034006 PMCID: PMC5053661 DOI: 10.18632/oncotarget.8443] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/11/2015] [Accepted: 03/17/2016] [Indexed: 01/24/2023] Open
Abstract
Human β-defensin-3 (hBD3) is an epithelial cell-derived innate immune regulatory molecule overexpressed in oral dysplastic lesions and fosters a tumor-promoting microenvironment. Expression of hBD3 is induced by the epidermal growth factor receptor signaling pathway. Here we describe a novel pathway through which the high-risk human papillomavirus type-16 (HPV-16) oncoprotein E6 induces hBD3 expression in mucosal keratinocytes. Ablation of E6 by siRNA induces the tumor suppressor p53 and diminishes hBD3 in HPV-16 positive CaSki cervical cancer cells and UM-SCC-104 head and neck cancer cells. Malignant cells in HPV-16-associated oropharyngeal cancer overexpress hBD3. HPV-16 E6 induces hBD3 mRNA expression, peptide production and gene promoter activity in mucosal keratinocytes. Reduction of cellular levels of p53 stimulates hBD3 expression, while activation of p53 by doxorubicin inhibits its expression in primary oral keratinocytes and CaSki cells, suggesting that p53 represses hBD3 expression. A p53 binding site in the hBD3 gene promoter has been identified by using electrophoretic mobility shift assays and chromatin immunoprecipitation (ChIP). In addition, the p63 protein isoform ΔNp63α, but not TAp63, stimulated transactivation of the hBD3 gene and was co-expressed with hBD3 in head and neck cancer specimens. Therefore, high-risk HPV E6 oncoproteins may stimulate hBD3 expression in tumor cells to facilitate tumorigenesis of HPV-associated head and neck cancer.
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Affiliation(s)
- Twishasri DasGupta
- Department of Biological Sciences, Case Western Reserve University School of Dental Medicine, Cleveland, OH, USA
| | - Emeka I Nweze
- Department of Biological Sciences, Case Western Reserve University School of Dental Medicine, Cleveland, OH, USA.,Present Address: University of Nigeria, Nsukka, Nigera
| | - Hong Yue
- Department of Biological Sciences, Case Western Reserve University School of Dental Medicine, Cleveland, OH, USA
| | - Liming Wang
- Center for Molecular Cancer Diagnosis Inc., Twinsburg, OH, USA
| | - Jessica Jin
- Human Developmental and Regenerative Biology, Harvard University, Cambridge, MA, USA
| | - Santosh K Ghosh
- Department of Biological Sciences, Case Western Reserve University School of Dental Medicine, Cleveland, OH, USA
| | - Hameem I Kawsar
- Department of Biological Sciences, Case Western Reserve University School of Dental Medicine, Cleveland, OH, USA.,Present Address: St. Luke's Hospital, Chesterfield, MO, USA
| | - Chad Zender
- Department of Otolaryngology-Head & Neck Surgery, University Hospitals Case Medical Center, Case Western Reserve University School of Medicine, Cleveland, OH, USA
| | - Elliot J Androphy
- Department of Dermatology, Indiana University School of Medicine, Indianapolis, IN, USA
| | - Aaron Weinberg
- Department of Biological Sciences, Case Western Reserve University School of Dental Medicine, Cleveland, OH, USA
| | - Thomas S McCormick
- Department of Dermatology, University Hospitals Case Medical Center, Case Western Reserve University School of Medicine, Cleveland, OH, USA
| | - Ge Jin
- Department of Biological Sciences, Case Western Reserve University School of Dental Medicine, Cleveland, OH, USA
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47
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Pinatti LM, Walline HM, Carey TE. Human Papillomavirus Genome Integration and Head and Neck Cancer. J Dent Res 2017; 97:691-700. [PMID: 29227715 DOI: 10.1177/0022034517744213] [Citation(s) in RCA: 42] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022] Open
Abstract
We conducted a critical review of human papillomavirus (HPV) integration into the host genome in oral/oropharyngeal cancer, reviewed the literature for HPV-induced cancers, and obtained current data for HPV-related oral and oropharyngeal cancers. In addition, we performed studies to identify HPV integration sites and the relationship of integration to viral-host fusion transcripts and whether integration is required for HPV-associated oncogenesis. Viral integration of HPV into the host genome is not required for the viral life cycle and might not be necessary for cellular transformation, yet HPV integration is frequently reported in cervical and head and neck cancer specimens. Studies of large numbers of early cervical lesions revealed frequent viral integration into gene-poor regions of the host genome with comparatively rare integration into cellular genes, suggesting that integration is a stochastic event and that site of integration may be largely a function of chance. However, more recent studies of head and neck squamous cell carcinomas (HNSCCs) suggest that integration may represent an additional oncogenic mechanism through direct effects on cancer-related gene expression and generation of hybrid viral-host fusion transcripts. In HNSCC cell lines as well as primary tumors, integration into cancer-related genes leading to gene disruption has been reported. The studies have shown that integration-induced altered gene expression may be associated with tumor recurrence. Evidence from several studies indicates that viral integration into genic regions is accompanied by local amplification, increased expression in some cases, interruption of gene expression, and likely additional oncogenic effects. Similarly, reported examples of viral integration near microRNAs suggest that altered expression of these regulatory molecules may also contribute to oncogenesis. Future work is indicated to identify the mechanisms of these events on cancer cell behavior.
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Affiliation(s)
- L M Pinatti
- 1 Cancer Biology Program, Program in the Biomedical Sciences, Rackham Graduate School, University of Michigan, Ann Arbor, MI, USA.,2 Department of Otolaryngology/Head and Neck Surgery, University of Michigan, Ann Arbor, MI, USA
| | - H M Walline
- 2 Department of Otolaryngology/Head and Neck Surgery, University of Michigan, Ann Arbor, MI, USA
| | - T E Carey
- 2 Department of Otolaryngology/Head and Neck Surgery, University of Michigan, Ann Arbor, MI, USA
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48
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Kohli S, Bhardwaj A, Kumari R, Das S. SIRT6 Is a Target of Regulation by UBE3A That Contributes to Liver Tumorigenesis in an ANXA2-Dependent Manner. Cancer Res 2017; 78:645-658. [PMID: 29217762 DOI: 10.1158/0008-5472.can-17-1692] [Citation(s) in RCA: 29] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/08/2017] [Revised: 09/29/2017] [Accepted: 11/29/2017] [Indexed: 01/23/2023]
Abstract
UBE3A is an E3 ubiquitin ligase well known for its role in the proteasomal degradation of p53 in human papillomavirus (HPV)-associated cancers. Here we report that UBE3A ubiquitylates and triggers degradation of the tumor-suppressive sirtuin SIRT6 in hepatocellular carcinoma. UBE3A ubiquitylated the highly conserved Lys160 residue on SIRT6. FOXO1-mediated transcriptional repression of UBE3A was sufficient to stabilize SIRT6 and to epigenetically repress ANXA2, a key mediator of UBE3A oncogenic function. Thus, UBE3A-mediated SIRT6 degradation promoted the proliferative capacity, migration potential, and invasiveness of cells. In mouse models of hepatocellular carcinoma, SIRT6 downregulation and consequent induction of ANXA2 were critical for UBE3A-mediated tumorigenesis. Furthermore, in clinical specimens, increased UBE3A levels correlated with reduced SIRT6 levels and elevated ANXA2 levels in increasing tumor grades. Overall, our findings show how the tumor suppressor SIRT6 is regulated in hepatocellular carcinoma and establish the mechanism underlying UBE3A-mediated tumorigenesis in this disease.Significance: These findings provide mechanistic insights into regulation of the tumor suppressive sirtuin SIRT6 and its implications for the development of hepatocellular carcinoma. Cancer Res; 78(3); 645-58. ©2017 AACR.
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Affiliation(s)
- Saishruti Kohli
- Molecular Oncology Laboratory, National Institute of Immunology, New Delhi, India
| | - Abhishek Bhardwaj
- Molecular Oncology Laboratory, National Institute of Immunology, New Delhi, India
| | - Richa Kumari
- Molecular Oncology Laboratory, National Institute of Immunology, New Delhi, India
| | - Sanjeev Das
- Molecular Oncology Laboratory, National Institute of Immunology, New Delhi, India.
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49
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Chhabra S, Kumar Y, Thacker G, Kapoor I, Lochab S, Sanyal S, Bhatt MLB, Chattopadhyay N, Trivedi AK. E6AP inhibits G-CSFR turnover and functions by promoting its ubiquitin-dependent proteasome degradation. BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH 2017; 1864:1545-1553. [PMID: 28578910 DOI: 10.1016/j.bbamcr.2017.05.026] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/21/2017] [Revised: 05/14/2017] [Accepted: 05/31/2017] [Indexed: 01/01/2023]
Abstract
Granulocyte colony-stimulating factor receptor (G-CSFR) plays a crucial role in regulating myeloid cell survival, proliferation, and neutrophilic granulocyte precursor cells maturation. Previously, we demonstrated that Fbw7α negatively regulates G-CSFR and its downstream signaling through ubiquitin-proteasome mediated degradation. However, whether additional ubiquitin ligases for G-CSFR exist is not known. Identifying multiple E3 ubiquitin ligases for G-CSFR shall improve our understanding of activation and subsequent attenuation of G-CSFR signaling required for differentiation and proliferation. Here, for the first time we demonstrate that E6 associated protein (E6AP), an E3 ubiquitin ligase physically associates with G-CSFR and targets it for ubiquitin-mediated proteasome degradation and thereby attenuates its functions. We further show that E6AP promoted G-CSFR degradation leads to reduced phosphorylation of signal transducer and activator of transcription 3 (STAT3) which is required for G-CSF dependent granulocytic differentiation. More importantly, our finding shows that E6AP also targets mutant form of G-SCFR (G-CSFR-T718), frequently observed in severe congenital neutropenia (SCN) patients that very often culminate to AML, however, at a quite slower rate than wild type G-CSFR. In addition, our data showed that knockdown of E6AP restores G-CSFR and its signaling thereby promoting granulocytic differentiation. Collectively, our data demonstrates that E6AP facilitates ubiquitination and subsequent degradation of G-CSFR leading to attenuation of its downstream signaling and inhibition of granulocytic differentiation.
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Affiliation(s)
- Stuti Chhabra
- Biochemistry Division, CSIR-Central Drug Research Institute, Sector-10, Jankipuram Extension, Sitapur Road, Lucknow, 226031, UP, India
| | - Yogesh Kumar
- Biochemistry Division, CSIR-Central Drug Research Institute, Sector-10, Jankipuram Extension, Sitapur Road, Lucknow, 226031, UP, India
| | - Gatha Thacker
- Biochemistry Division, CSIR-Central Drug Research Institute, Sector-10, Jankipuram Extension, Sitapur Road, Lucknow, 226031, UP, India
| | - Isha Kapoor
- Biochemistry Division, CSIR-Central Drug Research Institute, Sector-10, Jankipuram Extension, Sitapur Road, Lucknow, 226031, UP, India
| | - Savita Lochab
- Biochemistry Division, CSIR-Central Drug Research Institute, Sector-10, Jankipuram Extension, Sitapur Road, Lucknow, 226031, UP, India
| | - Sabyasachi Sanyal
- Biochemistry Division, CSIR-Central Drug Research Institute, Sector-10, Jankipuram Extension, Sitapur Road, Lucknow, 226031, UP, India
| | - Madan L B Bhatt
- Department of Radiotherapy, King George's Medical University, Lucknow, UP, India
| | - Naibedya Chattopadhyay
- Division of Endocrinology, CSIR-Central Drug Research Institute (CSIR-CDRI), Sector-10, Jankipuram Extension, Lucknow, 226031, UP, India
| | - Arun Kumar Trivedi
- Biochemistry Division, CSIR-Central Drug Research Institute, Sector-10, Jankipuram Extension, Sitapur Road, Lucknow, 226031, UP, India.
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50
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Piao S, Pei HZ, Huang B, Baek SH. Ovarian tumor domain-containing protein 1 deubiquitinates and stabilizes p53. Cell Signal 2017; 33:22-29. [DOI: 10.1016/j.cellsig.2017.02.011] [Citation(s) in RCA: 31] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2016] [Revised: 02/10/2017] [Accepted: 02/10/2017] [Indexed: 12/22/2022]
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