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Peng TW, Ma QF, Li J, Wang X, Zhang CH, Ma J, Li JY, Wang W, Zhu CL, Liu XH. HBV promotes its replication by up-regulating RAD51C gene expression. Sci Rep 2024; 14:2607. [PMID: 38297111 PMCID: PMC10831117 DOI: 10.1038/s41598-024-53047-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/30/2023] [Accepted: 01/27/2024] [Indexed: 02/02/2024] Open
Abstract
Chronic hepatitis B virus (HBV) infection is a major cause of hepatocellular carcinoma (HCC), pegylated-interferon-α(PEG-IFNα) and long-term nucleos(t)ide analogs (NUCs) are mainly drugs used to treat HBV infection, but the effectiveness is unsatisfactory in different populations, the exploration of novel therapeutic approaches is necessary. RAD51C is associated with DNA damage repair and plays an important role in the development and progression of tumors. Early cDNA microarray results showed that RAD51C expression was significantly increased in HBV-infected HCC cells, however, the relationship between HBV infection and abnormal expression of RAD51C has not been reported. Therefore, we conducted RT-PCR, western blot, Co-immunoprecipitation(Co-IP), and immunofluorescence(IF) to detect HBV-RAD51C interaction in RAD51C overexpression or interfering HCC cells. Our results showed that RAD51C and HBV X protein(HBX) produced a direct interaction in the nucleus, the HBV infection of HCC cells promoted RAD51C expression, and the increased expression of RAD51C promoted HBV replication. This indicated that RAD51C is closely related to the occurrence and development of HCC caused by HBV infection, and may bring a breakthrough in the the prevention and treatment study of HCC.
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Affiliation(s)
- Ting-Wei Peng
- Department of Clinical Laboratory, Shanghai Gongli Hospital, the Second Military Medical University, Shanghai, 200135, China
| | - Qing-Feng Ma
- Department of Clinical Laboratory, Liyuan Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430077, China
| | - Jie Li
- China Medical Tribune, Beijing, 100009, China
| | - Xue Wang
- Department of Clinical Laboratory, Shanghai Gongli Hospital, the Second Military Medical University, Shanghai, 200135, China
| | - Cong-Hui Zhang
- Department of Clinical Laboratory, Shanghai Gongli Hospital, the Second Military Medical University, Shanghai, 200135, China
| | - Junwen Ma
- Department of Clinical Laboratory, Liyuan Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430077, China
| | - Jun-Yi Li
- Department of Clinical Laboratory, Liyuan Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430077, China
| | - Wei Wang
- Department of Clinical Laboratory, Wuhan Fourth Hospital, Wuhan, 430034, China.
| | - Cheng-Liang Zhu
- Department of Clinical Laboratory, Renmin Hospital of Wuhan University, Wuhan, 430060, China.
| | - Xing-Hui Liu
- Department of Clinical Laboratory, Shanghai Gongli Hospital, the Second Military Medical University, Shanghai, 200135, China.
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2
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Anwaar A, Varma AK, Baruah R. In Silico-Based Structural Evaluation to Categorize the Pathogenicity of Mutations Identified in the RAD Class of Proteins. ACS OMEGA 2023; 8:10266-10277. [PMID: 36969410 PMCID: PMC10034773 DOI: 10.1021/acsomega.2c07802] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 12/07/2022] [Accepted: 02/10/2023] [Indexed: 06/18/2023]
Abstract
RAD genes, known as double-strand break repair proteins, play a major role in maintaining the genomic integrity of a cell by carrying out essential DNA repair functions via double-strand break repair pathways. Mutations in the RAD class of proteins show high susceptibility to breast and ovarian cancers; however, adequate research on the mutations identified in these genes has not been extensively reported for their deleterious effects. Changes in the folding pattern of RAD proteins play an important role in protein-protein interactions and also functions. Missense mutations identified from four cancer databases, cBioPortal, COSMIC, ClinVar, and gnomAD, cause aberrant conformations, which may lead to faulty DNA repair mechanisms. It is therefore necessary to evaluate the effects of pathogenic mutations of RAD proteins and their subsequent role in breast and ovarian cancers. In this study, we have used eight computational prediction servers to analyze pathogenic mutations and understand their effects on the protein structure and function. A total of 5122 missense mutations were identified from four different cancer databases, of which 1165 were predicted to be pathogenic using at least five pathogenicity prediction servers. These mutations were characterized as high-risk mutations based on their location in the conserved domains and subsequently subjected to structural stability characterization. The mutations included in the present study were selected from clinically relevant mutants in breast cancer pedigrees. Comparative folding patterns and intra-atomic interaction results showed alterations in the structural behavior of RAD proteins, specifically RAD51C triggered by mutations G125V and L138F and RAD51D triggered by mutations S207L and E233G.
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Affiliation(s)
- Aaliya Anwaar
- Advanced
Centre for Treatment, Research and Education in Cancer, Kharghar, Navi Mumbai 410210, Maharashtra, India
| | - Ashok K. Varma
- Advanced
Centre for Treatment, Research and Education in Cancer, Kharghar, Navi Mumbai 410210, Maharashtra, India
- Homi
Bhabha National Institute, Training School Complex, Anushaktinagar, Mumbai 400094, Maharashtra, India
| | - Reshita Baruah
- Advanced
Centre for Treatment, Research and Education in Cancer, Kharghar, Navi Mumbai 410210, Maharashtra, India
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Rawle DJ, Le TT, Dumenil T, Bishop C, Yan K, Nakayama E, Bird PI, Suhrbier A. Widespread discrepancy in Nnt genotypes and genetic backgrounds complicates granzyme A and other knockout mouse studies. eLife 2022; 11:e70207. [PMID: 35119362 PMCID: PMC8816380 DOI: 10.7554/elife.70207] [Citation(s) in RCA: 14] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/10/2021] [Accepted: 01/10/2022] [Indexed: 02/06/2023] Open
Abstract
Granzyme A (GZMA) is a serine protease secreted by cytotoxic lymphocytes, with Gzma-/- mouse studies having informed our understanding of GZMA's physiological function. We show herein that Gzma-/- mice have a mixed C57BL/6J and C57BL/6N genetic background and retain the full-length nicotinamide nucleotide transhydrogenase (Nnt) gene, whereas Nnt is truncated in C57BL/6J mice. Chikungunya viral arthritis was substantially ameliorated in Gzma-/- mice; however, the presence of Nnt and the C57BL/6N background, rather than loss of GZMA expression, was responsible for this phenotype. A new CRISPR active site mutant C57BL/6J GzmaS211A mouse provided the first insights into GZMA's bioactivity free of background issues, with circulating proteolytically active GZMA promoting immune-stimulating and pro-inflammatory signatures. Remarkably, k-mer mining of the Sequence Read Archive illustrated that ≈27% of Run Accessions and ≈38% of BioProjects listing C57BL/6J as the mouse strain had Nnt sequencing reads inconsistent with a C57BL/6J genetic background. Nnt and C57BL/6N background issues have clearly complicated our understanding of GZMA and may similarly have influenced studies across a broad range of fields.
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Affiliation(s)
- Daniel J Rawle
- QIMR Berghofer Medical Research InstituteBrisbaneAustralia
| | - Thuy T Le
- QIMR Berghofer Medical Research InstituteBrisbaneAustralia
| | - Troy Dumenil
- QIMR Berghofer Medical Research InstituteBrisbaneAustralia
| | - Cameron Bishop
- QIMR Berghofer Medical Research InstituteBrisbaneAustralia
| | - Kexin Yan
- QIMR Berghofer Medical Research InstituteBrisbaneAustralia
| | - Eri Nakayama
- QIMR Berghofer Medical Research InstituteBrisbaneAustralia
- Department of Virology I, National Institute of Infectious DiseasesTokyoJapan
| | - Phillip I Bird
- Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute, Monash UniversityMelbourneAustralia
| | - Andreas Suhrbier
- QIMR Berghofer Medical Research InstituteBrisbaneAustralia
- Australian Infectious Disease Research Centre, GVN Center of ExcellenceBrisbaneAustralia
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Transcriptional Differences in Lipid-Metabolizing Enzymes in Murine Sebocytes Derived from Sebaceous Glands of the Skin and Preputial Glands. Int J Mol Sci 2021; 22:ijms222111631. [PMID: 34769061 PMCID: PMC8584257 DOI: 10.3390/ijms222111631] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/05/2021] [Revised: 10/22/2021] [Accepted: 10/25/2021] [Indexed: 12/18/2022] Open
Abstract
Sebaceous glands are adnexal structures, which critically contribute to skin homeostasis and the establishment of a functional epidermal barrier. Sebocytes, the main cell population found within the sebaceous glands, are highly specialized lipid-producing cells. Sebaceous gland-resembling tissue structures are also found in male rodents in the form of preputial glands. Similar to sebaceous glands, they are composed of lipid-specialized sebocytes. Due to a lack of adequate organ culture models for skin sebaceous glands and the fact that preputial glands are much larger and easier to handle, previous studies used preputial glands as a model for skin sebaceous glands. Here, we compared both types of sebocytes, using a single-cell RNA sequencing approach, to unravel potential similarities and differences between the two sebocyte populations. In spite of common gene expression patterns due to general lipid-producing properties, we found significant differences in the expression levels of genes encoding enzymes involved in the biogenesis of specialized lipid classes. Specifically, genes critically involved in the mevalonate pathway, including squalene synthase, as well as the sphingolipid salvage pathway, such as ceramide synthase, (acid) sphingomyelinase or acid and alkaline ceramidases, were significantly less expressed by preputial gland sebocytes. Together, our data revealed tissue-specific sebocyte populations, indicating major developmental, functional as well as biosynthetic differences between both glands. The use of preputial glands as a surrogate model to study skin sebaceous glands is therefore limited, and major differences between both glands need to be carefully considered before planning an experiment.
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Li H, Ma Z, Che Z, Li Q, Fan J, Zhou Z, Wu Y, Jin Y, Liang P, Che X. Comprehensive role of prostate-specific antigen identified with proteomic analysis in prostate cancer. J Cell Mol Med 2020; 24:10202-10215. [PMID: 33107155 PMCID: PMC7520270 DOI: 10.1111/jcmm.15634] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/26/2020] [Accepted: 06/25/2020] [Indexed: 12/11/2022] Open
Abstract
Current treatments including androgen deprivation fail to prevent prostate cancer (PrCa) from progressing to castration-resistant PrCa (CRPC). Accumulating evidence highlights the relevance of prostate-specific antigen (PSA) in the development and progression of PrCa. The underlying mechanism whereby PSA functions in PrCa, however, has yet been elucidated. We demonstrated that PSA knockdown attenuated tumorigenesis and metastasis of PrCa C4-2 cells in vitro and in vivo, whereas promoted the apoptosis in vitro. To illuminate the comprehensive role of PSA in PrCa, we performed an isobaric tag for relative and absolute quantitation (iTRAQ)-based proteomic analysis to explore the proteomic change induced by PSA knockdown. Among 121 differentially expressed proteins, 67 proteins were up-regulated, while 54 proteins down-regulated. Bioinformatics analysis was used to explore the mechanism through which PSA exerts influence on PrCa. Protein-protein interaction analysis showed that PSA may mediate POTEF, EPHA3, RAD51C, HPGD and MCM4 to promote the initiation and progression of PrCa. We confirmed that PSA knockdown induced the up-regulation of MCM4 and RAD51C, while it down-regulated POTEF and EPHA3; meanwhile, MCM4 was higher in PrCa para-cancerous tissue than in cancerous tissue, suggesting that PSA may facilitate the tumorigenesis by mediating MCM4. Our findings suggest that PSA plays a comprehensive role in the development and progression of PrCa.
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Affiliation(s)
- Haoyong Li
- Department of Urology, Renmin Hospital of Wuhan University, Wuhan, China
| | - Zhe Ma
- Department of Urology, the First Affiliated Hospital of Hainan Medical University, Haikou, China
| | - Zhifei Che
- Department of Urology, the First Affiliated Hospital of Hainan Medical University, Haikou, China
| | - Qi Li
- Department of Urology, the First Affiliated Hospital of Hainan Medical University, Haikou, China
| | - Jinfeng Fan
- Department of Urology, the First Affiliated Hospital of Hainan Medical University, Haikou, China
| | - Zhiyan Zhou
- Department of Urology, the First Affiliated Hospital of Hainan Medical University, Haikou, China
| | - Yaoxi Wu
- Department of Urology, the First Affiliated Hospital of Hainan Medical University, Haikou, China
| | - Yingxia Jin
- Central Laboratory, Renmin Hospital of Wuhan University, Wuhan, China
| | - Peiyu Liang
- Department of Urology, the First Affiliated Hospital of Hainan Medical University, Haikou, China
| | - Xianping Che
- Department of Urology, The Second Affiliated Hospital of Hainan Medical University, Haikou, China
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6
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Ehrmann C, Schneider MR. Genetically modified laboratory mice with sebaceous glands abnormalities. Cell Mol Life Sci 2016; 73:4623-4642. [PMID: 27457558 PMCID: PMC11108334 DOI: 10.1007/s00018-016-2312-0] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/18/2016] [Revised: 07/12/2016] [Accepted: 07/19/2016] [Indexed: 12/19/2022]
Abstract
Sebaceous glands (SG) are exocrine glands that release their product by holocrine secretion, meaning that the whole cell becomes a secretion following disruption of the membrane. SG may be found in association with a hair follicle, forming the pilosebaceous unit, or as modified SG at different body sites such as the eyelids (Meibomian glands) or the preputial glands. Depending on their location, SG fulfill a number of functions, including protection of the skin and fur, thermoregulation, formation of the tear lipid film, and pheromone-based communication. Accordingly, SG abnormalities are associated with several diseases such as acne, cicatricial alopecia, and dry eye disease. An increasing number of genetically modified laboratory mouse lines develop SG abnormalities, and their study may provide important clues regarding the molecular pathways regulating SG development, physiology, and pathology. Here, we summarize in tabulated form the available mouse lines with SG abnormalities and, focusing on selected examples, discuss the insights they provide into SG biology and pathology. We hope this survey will become a helpful information source for researchers with a primary interest in SG but also as for researchers from unrelated fields that are unexpectedly confronted with a SG phenotype in newly generated mouse lines.
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Affiliation(s)
- Carmen Ehrmann
- Institute of Molecular Animal Breeding and Biotechnology, Gene Center, LMU Munich, Feodor-Lynen-Str. 25, 81377, Munich, Germany
| | - Marlon R Schneider
- Institute of Molecular Animal Breeding and Biotechnology, Gene Center, LMU Munich, Feodor-Lynen-Str. 25, 81377, Munich, Germany.
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Gu Y, Helenius M, Väänänen K, Bulanova D, Saarela J, Sokolenko A, Martens J, Imyanitov E, Kuznetsov S. BRCA1-deficient breast cancer cell lines are resistant to MEK inhibitors and show distinct sensitivities to 6-thioguanine. Sci Rep 2016; 6:28217. [PMID: 27313062 PMCID: PMC4911578 DOI: 10.1038/srep28217] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/13/2014] [Accepted: 06/01/2016] [Indexed: 12/27/2022] Open
Abstract
Germ-line or somatic inactivation of BRCA1 is a defining feature for a portion of human breast cancers. Here we evaluated the anti-proliferative activity of 198 FDA-approved and experimental drugs against four BRCA1-mutant (HCC1937, MDA-MB-436, SUM1315MO2, and SUM149PT) and four BRCA1-wild-type (MDA-MB-231, SUM229PE, MCF10A, and MCF7) breast cancer cell lines. We found that all BRCA1-mutant cell lines were insensitive to inhibitors of mitogen-activated protein kinase kinase 1 and 2 (MEK1/2) Selumetinib and Pimasertib in contrast to BRCA1-wildtype control cell lines. However, unexpectedly, only two BRCA1-mutant cell lines, HCC1937 and MDA-MB-436, were hypersensitive to a nucleotide analogue 6-thioguanine (6-TG). SUM149PT cells readily formed radiation-induced RAD51-positive nuclear foci indicating a functional homologous recombination, which may explain their resistance to 6-TG. However, the reason underlying 6-TG resistance of SUM1315MO2 cells remains unclear. Our data reveal a remarkable heterogeneity among BRCA1-mutant cell lines and provide a reference for future studies.
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Affiliation(s)
- Yuexi Gu
- Institute for Molecular Medicine Finland (FIMM), University of Helsinki, PO Box 20, FIN-00014, Helsinki, Finland
| | - Mikko Helenius
- Institute for Molecular Medicine Finland (FIMM), University of Helsinki, PO Box 20, FIN-00014, Helsinki, Finland
| | - Kristiina Väänänen
- Institute for Molecular Medicine Finland (FIMM), University of Helsinki, PO Box 20, FIN-00014, Helsinki, Finland.,Department of Biology, University of Eastern Finland, PO Box 111, FI-80101 Joensuu, Finland
| | - Daria Bulanova
- Institute for Molecular Medicine Finland (FIMM), University of Helsinki, PO Box 20, FIN-00014, Helsinki, Finland
| | - Jani Saarela
- Institute for Molecular Medicine Finland (FIMM), University of Helsinki, PO Box 20, FIN-00014, Helsinki, Finland
| | - Anna Sokolenko
- Department of Tumor Growth Biology, N.N. Petrov Institute of Oncology, St.-Petersburg 197758, Russia.,Department of Medical Genetics, St.-Petersburg Pediatric Medical University, St.-Petersburg 194100, Russia
| | - John Martens
- Erasmus University Medical Center, Daniel den Hoed Cancer Center, Department of Medical Oncology and Cancer Genomics Center, Dr. Molewaterplein 50, 3015 GE Rotterdam, the Netherlands
| | - Evgeny Imyanitov
- Department of Tumor Growth Biology, N.N. Petrov Institute of Oncology, St.-Petersburg 197758, Russia.,Department of Medical Genetics, St.-Petersburg Pediatric Medical University, St.-Petersburg 194100, Russia.,Department of Oncology, I.I. Mechnikov North-Western Medical University, St.-Petersburg 191015, Russia
| | - Sergey Kuznetsov
- Institute for Molecular Medicine Finland (FIMM), University of Helsinki, PO Box 20, FIN-00014, Helsinki, Finland
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Rad51c- and Trp53-double-mutant mouse model reveals common features of homologous recombination-deficient breast cancers. Oncogene 2016; 35:4601-10. [DOI: 10.1038/onc.2015.528] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/23/2015] [Revised: 11/14/2015] [Accepted: 12/18/2015] [Indexed: 01/15/2023]
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