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Li J, Lv L, Gao Y, Sun Y, Bai J, Wang X, Sun H, Jiang P. Tetraspanin CD81 serves as a functional entry factor for porcine circovirus type 2 infection. J Virol 2025; 99:e0140824. [PMID: 39745447 PMCID: PMC11853000 DOI: 10.1128/jvi.01408-24] [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] [Accepted: 12/06/2024] [Indexed: 02/26/2025] Open
Abstract
Porcine circovirus type 2 (PCV2) is the primary causative agent of porcine circovirus-associated disease, clinically resulting in immunosuppression and co-infections with other pathogens in infected pigs. The mechanism of PCV2 infection remains unclear. In this study, we firstly found that the tetraspanin CD81 in PK-15 cells interacts with PCV2 Cap protein by using virus overlay protein-binding assay combined with mass spectrometry. Knockdown of the CD81 significantly reduces the levels of the viral Cap mRNA and protein, and viral internalization in PK-15 cells. The critical interaction regions locate in the large extracellular loop (LEL) domain of CD81 and the CD loop region (82-91aa) of the Cap protein, and a polyclonal antibody against the CD81 LEL domain significantly inhibits PCV2 infection. The transmembrane proteoglycan Syndecan-1 interacts with both CD81 and PCV2 Cap, and co-operates with CD81 to promote PCV2 infection in PK-15 cells. Furthermore, CD81 facilitates RhoA activation and enhances the viral internalization and replication in PK-15 cells. It was concluded that the tetraspanin CD81 is a key host factor for PCV2 invasion into PK-15 cells, thus providing new insights into PCV2 life cycle and identifying a potential target for antiviral drug development.IMPORTANCEPorcine circovirus type 2 (PCV2), a significant economic pathogen in the swine industry, presents persistent challenges in its prevention and treatment. Despite extensive research, the mechanism of PCV2 invading host cells remains unclear. In this study, we found and identified a novel interaction between the tetraspanin CD81 and the viral Cap protein during the PCV2 invading PK-15 cells. The transmembrane proteoglycan Syndecan-1 and RhoA are involved in the infection process through the CD81. Moreover, this is the first time that the role of Syndecan-1 in the PCV2 infection process has been demonstrated. Also, a polyclonal antibody against the CD81 extracellular domain significantly inhibits PCV2 infection in PK-15 cells. It not only enriches our understanding of PCV2 life cycle but also offers new perspectives for the development of antiviral therapeutics against circovirus.
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Affiliation(s)
- Junshuo Li
- Key Laboratory of Animal Disease Diagnostics and Immunology, Ministry of Agriculture, MOE International Joint Collaborative Research Laboratory for Animal Health & Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, Jiangsu, China
| | - Lin Lv
- Key Laboratory of Animal Disease Diagnostics and Immunology, Ministry of Agriculture, MOE International Joint Collaborative Research Laboratory for Animal Health & Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, Jiangsu, China
| | - Yanni Gao
- Key Laboratory of Animal Disease Diagnostics and Immunology, Ministry of Agriculture, MOE International Joint Collaborative Research Laboratory for Animal Health & Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, Jiangsu, China
| | - Yangyang Sun
- Key Laboratory of Animal Disease Diagnostics and Immunology, Ministry of Agriculture, MOE International Joint Collaborative Research Laboratory for Animal Health & Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, Jiangsu, China
| | - Juan Bai
- Key Laboratory of Animal Disease Diagnostics and Immunology, Ministry of Agriculture, MOE International Joint Collaborative Research Laboratory for Animal Health & Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, Jiangsu, China
- Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou University, Yangzhou, Jiangsu, China
| | - Xianwei Wang
- Key Laboratory of Animal Disease Diagnostics and Immunology, Ministry of Agriculture, MOE International Joint Collaborative Research Laboratory for Animal Health & Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, Jiangsu, China
- Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou University, Yangzhou, Jiangsu, China
| | - Haifen Sun
- Key Laboratory of Animal Disease Diagnostics and Immunology, Ministry of Agriculture, MOE International Joint Collaborative Research Laboratory for Animal Health & Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, Jiangsu, China
| | - Ping Jiang
- Key Laboratory of Animal Disease Diagnostics and Immunology, Ministry of Agriculture, MOE International Joint Collaborative Research Laboratory for Animal Health & Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, Jiangsu, China
- Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou University, Yangzhou, Jiangsu, China
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Chuang YC, Ou JHJ. Hepatitis B virus entry, assembly, and egress. Microbiol Mol Biol Rev 2024; 88:e0001424. [PMID: 39440957 DOI: 10.1128/mmbr.00014-24] [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] [Indexed: 10/25/2024] Open
Abstract
SUMMARYHepatitis B virus (HBV) is an important human pathogen that chronically infects approximately 250 million people in the world, resulting in ~1 million deaths annually. This virus is a hepatotropic virus and can cause severe liver diseases including cirrhosis and hepatocellular carcinoma. The entry of HBV into hepatocytes is initiated by the interaction of its envelope proteins with its receptors. This is followed by the delivery of the viral nucleocapsid to the nucleus for the release of its genomic DNA and the transcription of viral RNAs. The assembly of the viral capsid particles may then take place in the nucleus or the cytoplasm and may involve cellular membranes. This is followed by the egress of the virus from infected cells. In recent years, significant research progresses had been made toward understanding the entry, the assembly, and the egress of HBV particles. In this review, we discuss the molecular pathways of these processes and compare them with those used by hepatitis delta virus and hepatitis C virus , two other hepatotropic viruses that are also enveloped. The understanding of these processes will help us to understand how HBV replicates and causes diseases, which will help to improve the treatments for HBV patients.
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Affiliation(s)
- Yu-Chen Chuang
- Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, California, USA
| | - J-H James Ou
- Department of Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Los Angeles, California, USA
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3
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Poddar S, Roy R, Kar P. The conformational dynamics of Hepatitis C Virus E2 glycoprotein with the increasing number of N-glycosylation unraveled by molecular dynamics simulations. J Biomol Struct Dyn 2024:1-16. [PMID: 38393644 DOI: 10.1080/07391102.2024.2319679] [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: 12/02/2023] [Accepted: 02/12/2024] [Indexed: 02/25/2024]
Abstract
The Hepatitis C Virus (HCV), responsible for causing hepatitis and a significant contributor to liver disorders, presents a challenge for treatment due to its high genetic variability. Despite efforts, there is still no effective medication available for this virus. One of the promising targets for drug development involves targeting glycoprotein E2. However, our understanding of the dynamic behavior of E2 and its associated glycans remains limited. In this study, we investigated the dynamic characteristics of E2 with varying degrees of glycosylation using all-atom molecular dynamics simulations. We also explored glycan's interactions with the protein and among themselves. An overall increase in correlation between the vital protein regions was observed with an increase in glycan number. The protein dynamics is followed by the analysis of glycan dynamics, where the flexibility of the individual glycans was analyzed in their free and bound state, which revealed a decrease in their fluctuation in some cases. Furthermore, we generated the free energy landscape of individual N-glycan linkages in both free and bound states and observed both increases and decreases in flexibility, which can be attributed to the formation and breakage of hydrogen bonds with amino acids. Finally, we found that for a high glycosylation system, glycans interact with glycoprotein and form hydrogen bonds among themselves. Moreover, the hydrogen bond profiles of a given glycan can vary when influenced by other glycans. In summary, our study provides valuable insights into the dynamics of the core region of HCV E2 glycoprotein and its associated glycans.
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Affiliation(s)
- Sayan Poddar
- Department of Biosciences and Biomedical Engineering, Indian Institute of Technology Indore, Indore, Madhya Pradesh, India
| | - Rajarshi Roy
- Department of Biosciences and Biomedical Engineering, Indian Institute of Technology Indore, Indore, Madhya Pradesh, India
| | - Parimal Kar
- Department of Biosciences and Biomedical Engineering, Indian Institute of Technology Indore, Indore, Madhya Pradesh, India
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So CW, Sourisseau M, Sarwar S, Evans MJ, Randall G. Roles of epidermal growth factor receptor, claudin-1 and occludin in multi-step entry of hepatitis C virus into polarized hepatoma spheroids. PLoS Pathog 2023; 19:e1011887. [PMID: 38157366 PMCID: PMC10756512 DOI: 10.1371/journal.ppat.1011887] [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: 08/17/2023] [Accepted: 12/06/2023] [Indexed: 01/03/2024] Open
Abstract
The multi-step process of hepatitis C virus (HCV) entry is facilitated by various host factors, including epidermal growth factor receptor (EGFR) and the tight junction proteins claudin-1 (CLDN1) and occludin (OCLN), which are thought to function at later stages of the HCV entry process. Using single particle imaging of HCV infection of polarized hepatoma spheroids, we observed that EGFR performs multiple functions in HCV entry, both phosphorylation-dependent and -independent. We previously observed, and in this study confirmed, that EGFR is not required for HCV migration to the tight junction. EGFR is required for the recruitment of clathrin to HCV in a phosphorylation-independent manner. EGFR phosphorylation is required for virion internalization at a stage following the recruitment of clathrin. HCV entry activates the RAF-MEK-ERK signaling pathway downstream of EGFR phosphorylation. This signaling pathway regulates the sorting and maturation of internalized HCV into APPL1- and EEA1-associated early endosomes, which form the site of virion uncoating. The tight junction proteins, CLDN1 and OCLN, function at two distinct stages of HCV entry. Despite its appreciated function as a "late receptor" in HCV entry, CLDN1 is required for efficient HCV virion accumulation at the tight junction. Huh-7.5 cells lacking CLDN1 accumulate HCV virions primarily at the initial basolateral surface. OCLN is required for the late stages of virion internalization. This study produced further insight into the unusually complex HCV endocytic process.
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Affiliation(s)
- Chui-Wa So
- Department of Microbiology, The University of Chicago, Chicago, Illinois, United States of America
| | - Marion Sourisseau
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America
| | - Shamila Sarwar
- Department of Microbiology, The University of Chicago, Chicago, Illinois, United States of America
| | - Matthew J. Evans
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America
| | - Glenn Randall
- Department of Microbiology, The University of Chicago, Chicago, Illinois, United States of America
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Carriquí-Madroñal B, Sheldon J, Duven M, Stegmann C, Cirksena K, Wyler E, Zapatero-Belinchón FJ, Vondran FWR, Gerold G. The matrix metalloproteinase ADAM10 supports hepatitis C virus entry and cell-to-cell spread via its sheddase activity. PLoS Pathog 2023; 19:e1011759. [PMID: 37967063 PMCID: PMC10650992 DOI: 10.1371/journal.ppat.1011759] [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: 03/14/2023] [Accepted: 10/16/2023] [Indexed: 11/17/2023] Open
Abstract
Hepatitis C virus (HCV) exploits the four entry factors CD81, scavenger receptor class B type I (SR-BI, also known as SCARB1), occludin, and claudin-1 as well as the co-factor epidermal growth factor receptor (EGFR) to infect human hepatocytes. Here, we report that the disintegrin and matrix metalloproteinase 10 (ADAM10) associates with CD81, SR-BI, and EGFR and acts as HCV host factor. Pharmacological inhibition, siRNA-mediated silencing and genetic ablation of ADAM10 reduced HCV infection. ADAM10 was dispensable for HCV replication but supported HCV entry and cell-to-cell spread. Substrates of the ADAM10 sheddase including epidermal growth factor (EGF) and E-cadherin, which activate EGFR family members, rescued HCV infection of ADAM10 knockout cells. ADAM10 did not influence infection with other enveloped RNA viruses such as alphaviruses and a common cold coronavirus. Collectively, our study reveals a critical role for the sheddase ADAM10 as a HCV host factor, contributing to EGFR family member transactivation and as a consequence to HCV uptake.
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Affiliation(s)
- Belén Carriquí-Madroñal
- Department of Biochemistry & Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hannover, Hanover, Germany
| | - Julie Sheldon
- Institute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, Hanover, Germany
| | - Mara Duven
- Department of Biochemistry & Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hannover, Hanover, Germany
| | - Cora Stegmann
- Department of Biochemistry & Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hannover, Hanover, Germany
| | - Karsten Cirksena
- Department of Biochemistry & Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hannover, Hanover, Germany
| | - Emanuel Wyler
- Max Delbrück Center for Molecular Medicine in the Helmholtz Association (MDC), Berlin Institute for Medical Systems Biology (BIMSB), Berlin, Germany
| | - Francisco J. Zapatero-Belinchón
- Department of Biochemistry & Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hannover, Hanover, Germany
- Gladstone Institutes, San Francisco, California, United States of America
| | - Florian W. R. Vondran
- Department of General, Visceral and Transplant Surgery, Regenerative Medicine and Experimental Surgery, Hannover Medical School, Hannover, Germany
- German Center for Infection Research Partner Site Hannover-Braunschweig Hannover, Germany
| | - Gisa Gerold
- Department of Biochemistry & Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hannover, Hanover, Germany
- Institute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, Hanover, Germany
- Wallenberg Centre for Molecular Medicine (WCMM), Umeå University, Umeå, Sweden
- Department of Clinical Microbiology, Virology, Umeå University, Umeå, Sweden
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Carriquí-Madroñal B, Lasswitz L, von Hahn T, Gerold G. Genetic and pharmacological perturbation of hepatitis-C virus entry. Curr Opin Virol 2023; 62:101362. [PMID: 37678113 DOI: 10.1016/j.coviro.2023.101362] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/02/2022] [Revised: 06/30/2023] [Accepted: 08/08/2023] [Indexed: 09/09/2023]
Abstract
Hepatitis-C virus (HCV) chronically infects 58 million individuals worldwide with variable disease outcome. While a subfraction of individuals exposed to the virus clear the infection, the majority develop chronic infection if untreated. Another subfraction of chronically ill proceeds to severe liver disease. The underlying causes of this interindividual variability include genetic polymorphisms in interferon genes. Here, we review available data on the influence of genetic or pharmacological perturbation of HCV host dependency factors on the clinically observed interindividual differences in disease outcome. We focus on host factors mediating virus entry into human liver cells. We assess available data on genetic variants of the major entry factors scavenger receptor class-B type I, CD81, claudin-1, and occludin as well as pharmacological perturbation of these entry factors. We review cell culture experimental and clinical cohort study data and conclude that entry factor perturbation may contribute to disease outcome of hepatitis C.
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Affiliation(s)
- Belén Carriquí-Madroñal
- Department of Biochemistry & Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hanover, Hanover, Germany; Institute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, Hanover, Germany
| | - Lisa Lasswitz
- Department of Biochemistry & Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hanover, Hanover, Germany; Institute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, Hanover, Germany
| | - Thomas von Hahn
- Department of Gastroenterology, Hepatology and Endocrinology, Hannover Medical School, 30625 Hannover, Germany; Department of Gastroenterology, Hepatology and Interventional Endoscopy, Asklepios Hospital Barmbek, Semmelweis University, Campus Hamburg, 22307 Hamburg, Germany
| | - Gisa Gerold
- Department of Biochemistry & Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hanover, Hanover, Germany; Institute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, Hanover, Germany; Wallenberg Centre for Molecular Medicine (WCMM), Umeå University, Umeå, Sweden; Department of Clinical Microbiology, Virology, Umeå University, Umeå, Sweden.
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7
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Ikram A, Rauff B, Alzahrani B, Awan FM, Obaid A, Naz A, Kakar SJ, Janjua HA. Integrated analysis to study the interplay between post-translational modifications (PTM) in hepatitis C virus proteins and hepatocellular carcinoma (HCC) development. Sci Rep 2022; 12:15648. [PMID: 36123370 PMCID: PMC9483894 DOI: 10.1038/s41598-022-19854-6] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/08/2021] [Accepted: 09/06/2022] [Indexed: 02/05/2023] Open
Abstract
Many PTMs dysregulation is known to be the major cause of many cancers including HCV induced HCC. PTMs of hepatitis C virus (HCV) regions NS3/4A, NS5A and NS5B are crucial for proper protein functions and replication that directly affect the generation of infectious virus particles and completion of its life cycle. In this study, we have performed comprehensive analysis of PTMs within HCV non-structural proteins (NS3/4A, NS5A and NS5B) through bioinformatics analysis to examine post-translational crosstalk between phosphorylation, palmitoylation, methylation, acetylation and ubiquitination sites in selected viral proteins. Our analysis has revealed many highly putative PTMs sites that are also conserved among major genotypes conferring the importance of these sites. We have also analysed viral 3D structures in their modified and unmodified forms to address extent and signatures of structural changes upon PTM. This study provides evidence that PTMs induce significant conformational changes and make viral proteins more stable. To find the potential role of PTMs in HCV induced HCC, docking analysis between selected viral proteins and p38-MAPK has been performed which also confirms their strong association with HCV induced HCC. The major findings proposed that PTMs at specific sites of HCV viral proteins could dysregulate specific pathways that cause the development of HCC.
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Affiliation(s)
- Aqsa Ikram
- Institute of Molecular Biology and Biotechnology (IMBB), University of Lahore (UOL), Lahore, Pakistan.
| | - Bisma Rauff
- Department of Biomedical Engineering, UET Lahore, Narowal campus, Narowal, Pakistan
| | - Badr Alzahrani
- Department of Clinical Laboratory Sciences, Jouf University, Sakaka, Saudi Arabia
| | - Faryal Mehwish Awan
- Department of Medical Lab Technology, University of Haripur (UOH), Haripur, Pakistan
| | - Ayesha Obaid
- Department of Medical Lab Technology, University of Haripur (UOH), Haripur, Pakistan
| | - Anam Naz
- Institute of Molecular Biology and Biotechnology (IMBB), University of Lahore (UOL), Lahore, Pakistan
| | - Salik Javed Kakar
- Atta Ur Rahman School of Applied Biosciences (ASAB), National University of Sciences and Technology (NUST), Islamabad, Pakistan
| | - Hussnain Ahmed Janjua
- Atta Ur Rahman School of Applied Biosciences (ASAB), National University of Sciences and Technology (NUST), Islamabad, Pakistan.
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Kim M, Kim M, Salloum S, Qian T, Wong LP, Xu M, Lee Y, Shroff SG, Sadreyev RI, Corey KE, Baumert TF, Hoshida Y, Chung RT. Atorvastatin favorably modulates a clinical hepatocellular carcinoma risk gene signature. Hepatol Commun 2022; 6:2581-2593. [PMID: 35712812 PMCID: PMC9426409 DOI: 10.1002/hep4.1991] [Citation(s) in RCA: 17] [Impact Index Per Article: 5.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/08/2021] [Revised: 04/11/2022] [Accepted: 04/15/2022] [Indexed: 11/22/2022] Open
Abstract
Lipophilic but not hydrophilic statins have been shown to be associated with reduced risk for hepatocellular carcinoma (HCC) in patients with chronic viral hepatitis. We investigated differential actions of lipophilic and hydrophilic statins and their ability to modulate a clinical prognostic liver signature (PLS) predicting HCC risk in patients with liver disease. Hepatitis C virus (HCV)-infected Huh7.5.1 cells, recently developed as a model to screen HCC chemopreventive agents, were treated with lipophilic statins (atorvastatin and simvastatin) and hydrophilic statins (rosuvastatin and pravastatin), and then analyzed by RNA sequencing and PLS. Lipophilic statins, particularly atorvastatin, more significantly suppressed the HCV-induced high-risk pattern of PLS and genes in YAP and AKT pathway implicated in fibrogenesis and carcinogenesis, compared with the hydrophilic statins. While atorvastatin inhibited YAP activation through the mevalonate pathway, the distinctive AKT inhibition of atorvastatin was mediated by stabilizing truncated retinoid X receptor alpha, which has been known to enhance AKT activation, representing a target for HCC chemoprevention. In addition, atorvastatin modulated the high-risk PLS in an in vitro model of nonalcoholic fatty liver disease (NAFLD). Conclusion: Atorvastatin distinctively inhibits YAP and AKT activation, which are biologically implicated in HCC development, and attenuates a high-risk PLS in an in vitro model of HCV infection and NAFLD. These findings suggest that atorvastatin is the most potent statin to reduce HCC risk in patients with viral and metabolic liver diseases.
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Affiliation(s)
- Myung‐Ho Kim
- Liver CenterGastrointestinal DivisionMassachusetts General HospitalBostonMassachusettsUSA
| | - Mi‐Young Kim
- Liver CenterGastrointestinal DivisionMassachusetts General HospitalBostonMassachusettsUSA
- Department of GastroenterologyCHA Bundang Medical CenterCHA University School of MedicineSeongnamSouth Korea
- Department of Gastroenterology, Chaum Life CenterCHA University School of MedicineSeoulSouth Korea
| | - Shadi Salloum
- Liver CenterGastrointestinal DivisionMassachusetts General HospitalBostonMassachusettsUSA
| | - Tongqi Qian
- Liver Tumor Translational Research ProgramSimmons Comprehensive Cancer CenterDivision of Digestive and Liver DiseasesDepartment of Internal MedicineUniversity of Texas Southwestern Medical CenterDallasTexasUSA
| | - Lai Ping Wong
- Department of Molecular BiologyMassachusetts General HospitalBostonMassachusettsUSA
- Department of GeneticsHarvard Medical SchoolBostonMassachusettsUSA
| | - Min Xu
- Liver CenterGastrointestinal DivisionMassachusetts General HospitalBostonMassachusettsUSA
| | - Yoojin Lee
- Liver CenterGastrointestinal DivisionMassachusetts General HospitalBostonMassachusettsUSA
| | - Stuti G. Shroff
- Department of PathologyMassachusetts General Hospital and Harvard Medical SchoolBostonMassachusettsUSA
| | - Ruslan I. Sadreyev
- Department of Molecular BiologyMassachusetts General HospitalBostonMassachusettsUSA
- Department of PathologyMassachusetts General Hospital and Harvard Medical SchoolBostonMassachusettsUSA
| | - Kathleen E. Corey
- Liver CenterGastrointestinal DivisionMassachusetts General HospitalBostonMassachusettsUSA
| | - Thomas F. Baumert
- Institut National de la Santé et de la Recherche MédicaleU1110Institut de Recherche sur les Maladies Virales et HépatiquesStrasbourgFrance
- Pole Hepato‐digestif, IHUStrasbourg University HospitalsStrasbourgFrance
| | - Yujin Hoshida
- Liver Tumor Translational Research ProgramSimmons Comprehensive Cancer CenterDivision of Digestive and Liver DiseasesDepartment of Internal MedicineUniversity of Texas Southwestern Medical CenterDallasTexasUSA
| | - Raymond T. Chung
- Liver CenterGastrointestinal DivisionMassachusetts General HospitalBostonMassachusettsUSA
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9
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Barman RK, Mukhopadhyay A, Maulik U, Das S. A network biology approach to identify crucial host targets for COVID-19. Methods 2022; 203:108-115. [PMID: 35364279 PMCID: PMC8960288 DOI: 10.1016/j.ymeth.2022.03.016] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/19/2021] [Revised: 03/09/2022] [Accepted: 03/27/2022] [Indexed: 12/23/2022] Open
Abstract
The ongoing global pandemic of COVID-19, caused by SARS-CoV-2 has killed more than 5.9 million individuals out of ∼43 million confirmed infections. At present, several parts of the world are encountering the 3rd wave. Mass vaccination has been started in several countries but they are less likely to be broadly available for the current pandemic, repurposing of the existing drugs has drawn highest attention for an immediate solution. A recent publication has mapped the physical interactions of SARS-CoV-2 and human proteins by affinity-purification mass spectrometry (AP-MS) and identified 332 high-confidence SARS-CoV-2-human protein-protein interactions (PPIs). Here, we taken a network biology approach and constructed a human protein-protein interaction network (PPIN) with the above SARS-CoV-2 targeted proteins. We utilized a combination of essential network centrality measures and functional properties of the human proteins to identify the critical human targets of SARS-CoV-2. Four human proteins, namely PRKACA, RHOA, CDK5RAP2, and CEP250 have emerged as the best therapeutic targets, of which PRKACA and CEP250 were also found by another group as potential candidates for drug targets in COVID-19. We further found candidate drugs/compounds, such as guanosine triphosphate, remdesivir, adenosine monophosphate, MgATP, and H-89 dihydrochloride that bind the target human proteins. The urgency to prevent the spread of infection and the death of diseased individuals has prompted the search for agents from the pool of approved drugs to repurpose them for COVID-19. Our results indicate that host targeting therapy with the repurposed drugs may be a useful strategy for the treatment of SARS-CoV-2 infection.
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Affiliation(s)
- Ranjan Kumar Barman
- Division of Virology, ICMR-National Institute of Cholera and Enteric Diseases, Kolkata 700010, India; Department of Computer Science and Engineering, Jadavpur University, Kolkata 700032, India
| | - Anirban Mukhopadhyay
- Department of Computer Science and Engineering, University of Kalyani, Kalyani 741235, West Bengal, India
| | - Ujjwal Maulik
- Department of Computer Science and Engineering, Jadavpur University, Kolkata 700032, India
| | - Santasabuj Das
- Division of Clinical Medicine, ICMR-National Institute of Cholera and Enteric Diseases, Kolkata 700010, India; ICMR-National Institute of Occupational Health, Ahmedabad 380016, India.
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10
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Li HC, Yang CH, Lo SY. Cellular factors involved in the hepatitis C virus life cycle. World J Gastroenterol 2021; 27:4555-4581. [PMID: 34366623 PMCID: PMC8326260 DOI: 10.3748/wjg.v27.i28.4555] [Citation(s) in RCA: 13] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/27/2021] [Revised: 04/04/2021] [Accepted: 07/09/2021] [Indexed: 02/06/2023] Open
Abstract
The hepatitis C virus (HCV), an obligatory intracellular pathogen, highly depends on its host cells to propagate successfully. The HCV life cycle can be simply divided into several stages including viral entry, protein translation, RNA replication, viral assembly and release. Hundreds of cellular factors involved in the HCV life cycle have been identified over more than thirty years of research. Characterization of these cellular factors has provided extensive insight into HCV replication strategies. Some of these cellular factors are targets for anti-HCV therapies. In this review, we summarize the well-characterized and recently identified cellular factors functioning at each stage of the HCV life cycle.
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Affiliation(s)
- Hui-Chun Li
- Department of Biochemistry, Tzu Chi University, Hualien 970, Taiwan
| | - Chee-Hing Yang
- Department of Laboratory Medicine and Biotechnology, Tzu Chi University, Hualien 970, Taiwan
| | - Shih-Yen Lo
- Department of Laboratory Medicine and Biotechnology, Tzu Chi University, Hualien 970, Taiwan
- Department of Laboratory Medicine, Buddhist Tzu Chi General Hospital, Hualien 970, Taiwan
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Porcine Sapovirus-Induced Tight Junction Dissociation via Activation of RhoA/ROCK/MLC Signaling Pathway. J Virol 2021; 95:JVI.00051-21. [PMID: 33692204 PMCID: PMC8139687 DOI: 10.1128/jvi.00051-21] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
Tight junctions (TJs) are a major barrier and also an important portal of entry for different pathogens. Porcine sapovirus (PSaV) induces early disruption of the TJ integrity of polarized LLC-PK cells, allowing it to bind to the buried occludin co-receptors hidden beneath the TJs on the basolateral surface. However, the signaling pathways involved in the PSaV-induced TJ dissociation are not yet known. Here, we found that the RhoA/ROCK/MLC signaling pathway was activated in polarized LLC-PK cells during the early infection of PSaV Cowden strain in the presence of bile acid. Specific inhibitors of RhoA, ROCK, and MLC restored PSaV-induced reduction of transepithelial resistance, increase of paracellular flux, intracellular translocation of occludin, and lateral membrane lipid diffusion. Moreover, each inhibitor significantly reduced PSaV replication, as evidenced by a reduction in viral protein synthesis, genome copy number, and progeny viruses. The PKC/MLCK and RhoA/ROCK/MYPT signaling pathways, known to dissociate TJs, were not activated during early PSaV infection. Among the above signaling pathways, the RhoA/ROCK/MLC signaling pathway was only activated by PSaV in the absence of bile acid, and specific inhibitors of this signaling pathway restored early TJ dissociation. Our findings demonstrate that PSaV binding to cell surface receptors activates the RhoA/ROCK/MLC signaling pathway, which in turn disrupts TJ integrity via the contraction of the actomyosin ring. Our study contributes to understanding how PSaV enters the cells and will aid in developing efficient and affordable therapies against PSaV and other calicivirus infections.IMPORTANCEPorcine sapovirus (PSaV), one of the most important enteric pathogens, is known to disrupt tight junction (TJ) integrity to expose its buried co-receptor occludin in polarized LLC-PK cells. However, the cellular signaling pathways that facilitate TJ dissociation are not yet completely understood. Here, we demonstrate that early infection of PSaV in polarized LLC-PK cells in either the presence or absence of bile acids activates the RhoA/ROCK/MLC signaling pathway, whose inhibitors reverse the early PSaV infection-induced early dissociation of TJs and reduce PSaV replication. However, early PSaV infection did not activate the PKC/MLCK and RhoA/ROCK/MYPT signaling pathways, which are also known to dissociate TJs. This study provides a better understanding of the mechanism involved in early PSaV infection-induced disruption of TJs, which is important for controlling or preventing PSaV and other calicivirus infections.
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Velázquez-Moctezuma R, Augestad EH, Castelli M, Holmboe Olesen C, Clementi N, Clementi M, Mancini N, Prentoe J. Mechanisms of Hepatitis C Virus Escape from Vaccine-Relevant Neutralizing Antibodies. Vaccines (Basel) 2021; 9:291. [PMID: 33804732 PMCID: PMC8004074 DOI: 10.3390/vaccines9030291] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/24/2021] [Revised: 03/15/2021] [Accepted: 03/16/2021] [Indexed: 12/15/2022] Open
Abstract
Hepatitis C virus (HCV) is a major causative agent of acute and chronic hepatitis. It is estimated that 400,000 people die every year from chronic HCV infection, mostly from severe liver-related diseases such as cirrhosis and liver cancer. Although HCV was discovered more than 30 years ago, an efficient prophylactic vaccine is still missing. The HCV glycoprotein complex, E1/E2, is the principal target of neutralizing antibodies (NAbs) and, thus, is an attractive antigen for B-cell vaccine design. However, the high genetic variability of the virus necessitates the identification of conserved epitopes. Moreover, the high intrinsic mutational capacity of HCV allows the virus to continually escape broadly NAbs (bNAbs), which is likely to cause issues with vaccine-resistant variants. Several studies have assessed the barrier-to-resistance of vaccine-relevant bNAbs in vivo and in vitro. Interestingly, recent studies have suggested that escape substitutions can confer antibody resistance not only by direct modification of the epitope but indirectly through allosteric effects, which can be grouped based on the breadth of these effects on antibody susceptibility. In this review, we summarize the current understanding of HCV-specific NAbs, with a special focus on vaccine-relevant bNAbs and their targets. We highlight antibody escape studies pointing out the different methodologies and the escape mutations identified thus far. Finally, we analyze the antibody escape mechanisms of envelope protein escape substitutions and polymorphisms according to the most recent evidence in the HCV field. The accumulated knowledge in identifying bNAb epitopes as well as assessing barriers to resistance and elucidating relevant escape mechanisms may prove critical in the successful development of an HCV B-cell vaccine.
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Affiliation(s)
- Rodrigo Velázquez-Moctezuma
- Copenhagen Hepatitis C Program (CO-HEP), Department of Immunology and Microbiology, Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen, Denmark; (R.V.-M.); (E.H.A.); (C.H.O.)
- Department of Infectious Diseases, Hvidovre Hospital, 2650 Hvidovre, Denmark
| | - Elias H. Augestad
- Copenhagen Hepatitis C Program (CO-HEP), Department of Immunology and Microbiology, Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen, Denmark; (R.V.-M.); (E.H.A.); (C.H.O.)
- Department of Infectious Diseases, Hvidovre Hospital, 2650 Hvidovre, Denmark
| | - Matteo Castelli
- Laboratory of Microbiology and Virology, Università “Vita-Salute” San Raffaele, 20132 Milano, Italy; (M.C.); (N.C.); (M.C.); (N.M.)
| | - Christina Holmboe Olesen
- Copenhagen Hepatitis C Program (CO-HEP), Department of Immunology and Microbiology, Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen, Denmark; (R.V.-M.); (E.H.A.); (C.H.O.)
- Department of Infectious Diseases, Hvidovre Hospital, 2650 Hvidovre, Denmark
| | - Nicola Clementi
- Laboratory of Microbiology and Virology, Università “Vita-Salute” San Raffaele, 20132 Milano, Italy; (M.C.); (N.C.); (M.C.); (N.M.)
| | - Massimo Clementi
- Laboratory of Microbiology and Virology, Università “Vita-Salute” San Raffaele, 20132 Milano, Italy; (M.C.); (N.C.); (M.C.); (N.M.)
| | - Nicasio Mancini
- Laboratory of Microbiology and Virology, Università “Vita-Salute” San Raffaele, 20132 Milano, Italy; (M.C.); (N.C.); (M.C.); (N.M.)
| | - Jannick Prentoe
- Copenhagen Hepatitis C Program (CO-HEP), Department of Immunology and Microbiology, Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen, Denmark; (R.V.-M.); (E.H.A.); (C.H.O.)
- Department of Infectious Diseases, Hvidovre Hospital, 2650 Hvidovre, Denmark
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Investigating virus-host cell interactions: Comparative binding forces between hepatitis C virus-like particles and host cell receptors in 2D and 3D cell culture models. J Colloid Interface Sci 2021; 592:371-384. [PMID: 33677197 DOI: 10.1016/j.jcis.2021.02.067] [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: 11/24/2020] [Revised: 02/13/2021] [Accepted: 02/15/2021] [Indexed: 12/25/2022]
Abstract
Cell cultures have been successfully used to study hepatitis C virus (HCV) for many years. However, most work has been done using traditional, 2-dimensional (2D) cell cultures (cells grown as a monolayer in growth flasks or dishes). Studies have shown that when cells are grown suspended in an extra-cellular-matrix-like material, they develop into spherical, 'organoid' arrangements of cells (3D growth) that display distinct differences in morphological and functional characteristics compared to 2D cell cultures. In liver organoids, one key difference is the development of clearly differentiated apical and basolateral surfaces separated and maintained by cellular tight junctions. This phenomenon, termed polarity, is vital to normal barrier function of hepatocytes in vivo. It has also been shown that viruses, and virus-like particles, interact very differently with cells derived from 2D as compared to 3D cell cultures, bringing into question the usefulness of 2D cell cultures to study virus-host cell interactions. Here, we investigate differences in cellular architecture as a function of cell culture system, using confocal scanning laser microscopy, and determine differences in binding interactions between HCV virus-like particles (VLPs) and their cognate receptors in the different cell culture systems using atomic force microscopy (AFM). We generated organoid cultures that were polarized, as determined by localization of key apical and basolateral markers. We found that, while uptake of HCV VLPs by both 2D and 3D Huh7 cells was observed by flow cytometry, binding interactions between HCV VLPs and cells were measurable by AFM only on polarized cells. The work presented here adds to the growing body of research suggesting that polarized cell systems are more suitable for the study of HCV infection and dynamics than non-polarized systems.
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Alberione MP, Moeller R, Kirui J, Ginkel C, Doepke M, Ströh LJ, Machtens JP, Pietschmann T, Gerold G. Single-nucleotide variants in human CD81 influence hepatitis C virus infection of hepatoma cells. Med Microbiol Immunol 2020; 209:499-514. [PMID: 32322956 PMCID: PMC7176029 DOI: 10.1007/s00430-020-00675-1] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/20/2019] [Accepted: 04/06/2020] [Indexed: 02/07/2023]
Abstract
An estimated number of 71 million people are living with chronic hepatitis C virus (HCV) infection worldwide and 400,000 annual deaths are related to the infection. HCV entry into the hepatocytes is complex and involves several host factors. The tetraspanin human CD81 (hCD81) is one of the four essential entry factors and is composed of one large extracellular loop, one small extracellular loop, four transmembrane domains, one intracellular loop and two intracellular tails. The large extracellular loop interacts with the E2 glycoprotein of HCV. Regions outside the large extracellular loop (backbone) of hCD81 have a critical role in post-binding entry steps and determine susceptibility of hepatocytes to HCV. Here, we investigated the effect of five non-synonymous single-nucleotide variants in the backbone of hCD81 on HCV susceptibility. We generated cell lines that stably express the hCD81 variants and infected the cells using HCV pseudoparticles and cell culture-derived HCV. Our results show that all the tested hCD81 variants support HCV pseudoparticle entry with similar efficiency as wild-type hCD81. In contrast, variants A54V, V211M and M220I are less supportive to cell culture-derived HCV infection. This altered susceptibility is HCV genotype dependent and specifically affected the cell entry step. Our findings identify three hCD81 genetic variants that are impaired in their function as HCV host factors for specific viral genotypes. This study provides additional evidence that genetic host variation contributes to inter-individual differences in HCV infection and outcome.
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Affiliation(s)
- María Pía Alberione
- Institute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, Feodor-Lynen-Straße 7, 30625, Hannover, Germany
| | - Rebecca Moeller
- Institute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, Feodor-Lynen-Straße 7, 30625, Hannover, Germany
| | - Jared Kirui
- Institute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, Feodor-Lynen-Straße 7, 30625, Hannover, Germany
| | - Corinne Ginkel
- Institute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, Feodor-Lynen-Straße 7, 30625, Hannover, Germany
| | - Mandy Doepke
- Institute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, Feodor-Lynen-Straße 7, 30625, Hannover, Germany
| | - Luisa J Ströh
- Institute of Virology, Hannover Medical School, Hannover, Germany
| | - Jan-Philipp Machtens
- Institute of Biological Information Processing (IBI-1), Molekular- und Zellphysiologie, and JARA-HPC, Forschungszentrum Jülich, Jülich, Germany
- Institute of Clinical Pharmacology, RWTH Aachen University, Aachen, Germany
| | - Thomas Pietschmann
- Institute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, Feodor-Lynen-Straße 7, 30625, Hannover, Germany
| | - Gisa Gerold
- Institute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, Feodor-Lynen-Straße 7, 30625, Hannover, Germany.
- Department of Clinical Microbiology, Virology and Wallenberg Centre for Molecular Medicine (WCMM), Umeå University, Umeå, Sweden.
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Mailly L, Baumert TF. Hepatitis C virus infection and tight junction proteins: The ties that bind. BIOCHIMICA ET BIOPHYSICA ACTA-BIOMEMBRANES 2020; 1862:183296. [PMID: 32268133 DOI: 10.1016/j.bbamem.2020.183296] [Citation(s) in RCA: 20] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/13/2020] [Revised: 03/25/2020] [Accepted: 03/28/2020] [Indexed: 02/07/2023]
Abstract
The hepatitis C virus (HCV) is a major cause of liver diseases ranging from liver inflammation to advanced liver diseases like cirrhosis and hepatocellular carcinoma (HCC). HCV infection is restricted to the liver, and more specifically to hepatocytes, which represent around 80% of liver cells. The mechanism of HCV entry in human hepatocytes has been extensively investigated since the discovery of the virus 30 years ago. The entry mechanism is a multi-step process relying on several host factors including heparan sulfate proteoglycan (HSPG), low density lipoprotein receptor (LDLR), tetraspanin CD81, Scavenger Receptor class B type I (SR-BI), Epidermal Growth Factor Receptor (EGFR) and Niemann-Pick C1-like 1 (NPC1L1). Moreover, in order to establish a persistent infection, HCV entry is dependent on the presence of tight junction (TJ) proteins Claudin-1 (CLDN1) and Occludin (OCLN). In the liver, tight junction proteins play a role in architecture and homeostasis including sealing the apical pole of adjacent cells to form bile canaliculi and separating the basolateral domain drained by sinusoidal blood flow. In this review, we will highlight the role of liver tight junction proteins in HCV infection, and we will discuss the potential targeted therapeutic approaches to improve virus eradication.
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Affiliation(s)
- Laurent Mailly
- Université de Strasbourg, INSERM, UMR-S1110, Institut de Recherche sur les Maladies Virales et Hépatiques, F-67000 Strasbourg, France.
| | - Thomas F Baumert
- Université de Strasbourg, INSERM, UMR-S1110, Institut de Recherche sur les Maladies Virales et Hépatiques, F-67000 Strasbourg, France; Pôle Hépato-digestif, Hôpitaux Universitaires de Strasbourg, F-67000 Strasbourg, France; Institut Universitaire de France, F-75231 Paris, France.
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16
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Hepatitis C Virus Entry: An Intriguingly Complex and Highly Regulated Process. Int J Mol Sci 2020; 21:ijms21062091. [PMID: 32197477 PMCID: PMC7140000 DOI: 10.3390/ijms21062091] [Citation(s) in RCA: 16] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/27/2020] [Revised: 03/15/2020] [Accepted: 03/16/2020] [Indexed: 02/07/2023] Open
Abstract
Hepatitis C virus (HCV) is a major cause of chronic hepatitis and liver disease worldwide. Its tissue and species tropism are largely defined by the viral entry process that is required for subsequent productive viral infection and establishment of chronic infection. This review provides an overview of the viral and host factors involved in HCV entry into hepatocytes, summarizes our understanding of the molecular mechanisms governing this process and highlights the therapeutic potential of host-targeting entry inhibitors.
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17
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Kurniawan DW, Storm G, Prakash J, Bansal R. Role of spleen tyrosine kinase in liver diseases. World J Gastroenterol 2020; 26:1005-1019. [PMID: 32205992 PMCID: PMC7081001 DOI: 10.3748/wjg.v26.i10.1005] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/29/2019] [Revised: 01/14/2020] [Accepted: 02/28/2020] [Indexed: 02/06/2023] Open
Abstract
Spleen tyrosine kinase (SYK) is a non-receptor tyrosine kinase expressed in most hematopoietic cells and non-hematopoietic cells and play a crucial role in both immune and non-immune biological responses. SYK mediate diverse cellular responses via an immune-receptor tyrosine-based activation motifs (ITAMs)-dependent signalling pathways, ITAMs-independent and ITAMs-semi-dependent signalling pathways. In liver, SYK expression has been observed in parenchymal (hepatocytes) and non-parenchymal cells (hepatic stellate cells and Kupffer cells), and found to be positively correlated with the disease severity. The implication of SYK pathway has been reported in different liver diseases including liver fibrosis, viral hepatitis, alcoholic liver disease, non-alcoholic steatohepatitis and hepatocellular carcinoma. Antagonism of SYK pathway using kinase inhibitors have shown to attenuate the progression of liver diseases thereby suggesting SYK as a highly promising therapeutic target. This review summarizes the current understanding of SYK and its therapeutic implication in liver diseases.
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Affiliation(s)
- Dhadhang Wahyu Kurniawan
- Department of Biomaterials Science and Technology, Faculty of Science and Technology, Technical Medical Centre, University of Twente, Enschede 7500, the Netherlands
- Department of Pharmacy, Universitas Jenderal Soedirman, Purwokerto 53132, Indonesia
| | - Gert Storm
- Department of Biomaterials Science and Technology, Faculty of Science and Technology, Technical Medical Centre, University of Twente, Enschede 7500, the Netherlands
- Department of Pharmaceutics, University of Utrecht, Utrecht 3454, the Netherlands
| | - Jai Prakash
- Department of Biomaterials Science and Technology, Faculty of Science and Technology, Technical Medical Centre, University of Twente, Enschede 7500, the Netherlands
| | - Ruchi Bansal
- Department of Biomaterials Science and Technology, Faculty of Science and Technology, Technical Medical Centre, University of Twente, Enschede 7500, the Netherlands
- Department of Pharmacokinetics, Toxicology and Targeting, Groningen Research Institute of Pharmacy, University of Groningen, Enschede 7500, the Netherlands
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18
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Gerold G, Moeller R, Pietschmann T. Hepatitis C Virus Entry: Protein Interactions and Fusion Determinants Governing Productive Hepatocyte Invasion. Cold Spring Harb Perspect Med 2020; 10:cshperspect.a036830. [PMID: 31427285 DOI: 10.1101/cshperspect.a036830] [Citation(s) in RCA: 33] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Hepatitis C virus (HCV) entry is among the best-studied uptake processes for human pathogenic viruses. Uptake follows a spatially and temporally tightly controlled program. Numerous host factors including proteins, lipids, and glycans promote productive uptake of HCV particles into human liver cells. The virus initially attaches to surface proteoglycans, lipid receptors such as the scavenger receptor BI (SR-BI), and to the tetraspanin CD81. After lateral translocation of virions to tight junctions, claudin-1 (CLDN1) and occludin (OCLN) are essential for entry. Clathrin-mediated endocytosis engulfs HCV particles, which fuse with endosomal membranes after pH drop. Uncoating of the viral RNA genome in the cytoplasm completes the entry process. Here we systematically review and classify HCV entry factors by their mechanistic role, relevance, and level of evidence. Finally, we report on more recent knowledge on determinants of membrane fusion and close with an outlook on future implications of HCV entry research.
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Affiliation(s)
- Gisa Gerold
- TWINCORE, Center for Experimental and Clinical Infection Research, Institute for Experimental Virology, 30625 Hannover, Germany.,Department of Clinical Microbiology, Virology & Wallenberg Centre for Molecular Medicine (WCMM), Umeå University, 901 85 Umeå, Sweden
| | - Rebecca Moeller
- TWINCORE, Center for Experimental and Clinical Infection Research, Institute for Experimental Virology, 30625 Hannover, Germany
| | - Thomas Pietschmann
- TWINCORE, Center for Experimental and Clinical Infection Research, Institute for Experimental Virology, 30625 Hannover, Germany
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Pol S, Lagaye S. The remarkable history of the hepatitis C virus. Microbes Infect 2019; 21:263-270. [PMID: 31295571 DOI: 10.1016/j.micinf.2019.06.008] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/28/2019] [Revised: 03/25/2019] [Accepted: 04/01/2019] [Indexed: 12/23/2022]
Abstract
The infection with the hepatitis C virus (HCV) is an example of the translational research success. The reciprocal interactions between clinicians and scientists have allowed in 30 years the initiation of empirical treatments by interferon, the discovery of the virus, the development of serological and virological tools for diagnosis but also for prognosis (the non-invasive biochemical or morphological fibrosis tests, the predictors of the specific immune response including genetic IL28B polymorphisms). Finally, well-tolerated and effective treatments with oral antivirals inhibiting HCV non-structural viral proteins involved in viral replication have been marketed this last decade, allowing the cure of all infected subjects. HCV chronic infection, which is a public health issue, is a hepatic disease which may lead to a cirrhosis and an hepatocellular carcinoma (HCC) but also a systemic disease with extra-hepatic manifestations either associated with a cryoglobulinemic vasculitis or chronic inflammation. The HCV infection is the only chronic viral infection which may be cured: the so-called sustained virologic response, defined by undetectable HCV RNA 12 weeks after the end of the treatment, significantly reduces the risk of morbidity and mortality associated with hepatic and extra-hepatic manifestations which are mainly reversible. The history of HCV ends with the pangenotypic efficacy of the multiple combinations, easy to use for 8-12 weeks with one to three pills per day and little problems of tolerance. This explains the short 30 years from the virus discovery to the viral hepatitis elimination policy proposed by the World Health Organization (WHO) in 2016.
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Affiliation(s)
- Stanislas Pol
- Université Paris Descartes, Paris, France; Département d'Hépatologie, Hôpital Cochin, APHP, Paris, France; INSERM UMS-20, Institut Pasteur, Paris, France; Immunobiologie des Cellules Dendritiques, Institut Pasteur, Paris, France; INSERM U1223, Institut Pasteur, Paris, France.
| | - Sylvie Lagaye
- Immunobiologie des Cellules Dendritiques, Institut Pasteur, Paris, France; INSERM U1223, Institut Pasteur, Paris, France.
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Perez S, Kaspi A, Domovitz T, Davidovich A, Lavi-Itzkovitz A, Meirson T, Alison Holmes J, Dai CY, Huang CF, Chung RT, Nimer A, El-Osta A, Yaari G, Stemmer SM, Yu ML, Haviv I, Gal-Tanamy M. Hepatitis C virus leaves an epigenetic signature post cure of infection by direct-acting antivirals. PLoS Genet 2019; 15:e1008181. [PMID: 31216276 PMCID: PMC6602261 DOI: 10.1371/journal.pgen.1008181] [Citation(s) in RCA: 116] [Impact Index Per Article: 19.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/11/2018] [Revised: 07/01/2019] [Accepted: 05/08/2019] [Indexed: 02/06/2023] Open
Abstract
The increasing worldwide prevalence of Hepatocellular carcinoma (HCC), characterized by resistance to conventional chemotherapy, poor prognosis and eventually mortality, place it as a prime target for new modes of prevention and treatment. Hepatitis C Virus (HCV) is the predominant risk factor for HCC in the US and Europe. Multiple epidemiological studies showed that sustained virological responses (SVR) following treatment with the powerful direct acting antivirals (DAAs), which have replaced interferon-based regimes, do not eliminate tumor development. We aimed to identify an HCV-specific pathogenic mechanism that persists post SVR following DAAs treatment. We demonstrate that HCV infection induces genome-wide epigenetic changes by performing chromatin immunoprecipitation followed by next-generation sequencing (ChIP-seq) for histone post-translational modifications that are epigenetic markers for active and repressed chromatin. The changes in histone modifications correlate with reprogramed host gene expression and alter signaling pathways known to be associated with HCV life cycle and HCC. These epigenetic alterations require the presence of HCV RNA or/and expression of the viral proteins in the cells. Importantly, the epigenetic changes induced following infection persist as an "epigenetic signature" after virus eradication by DAAs treatment, as detected using in vitro HCV infection models. These observations led to the identification of an 8 gene signature that is associated with HCC development and demonstrate persistent epigenetic alterations in HCV infected and post SVR liver biopsy samples. The epigenetic signature was reverted in vitro by drugs that inhibit epigenetic modifying enzyme and by the EGFR inhibitor, Erlotinib. This epigenetic "scarring" of the genome, persisting following HCV eradication, suggest a novel mechanism for the persistent pathogenesis of HCV after its eradication by DAAs. Our study offers new avenues for prevention of the persistent oncogenic effects of chronic hepatitis infections using specific drugs to revert the epigenetic changes to the genome.
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Affiliation(s)
- Shira Perez
- Molecular Virology Lab, Azrieli Faculty of Medicine in the Galilee, Bar-Ilan University, Safed, Israel
- Cancer Personalized Medicine and Diagnostic Genomics Lab, Azrieli Faculty of Medicine in the Galilee, Bar-Ilan University, Safed, Israel
| | - Antony Kaspi
- Epigenetics in Human Health and Disease Laboratory, Department of Diabetes, Central Clinical School, Monash University, Melbourne, Australia
| | - Tom Domovitz
- Molecular Virology Lab, Azrieli Faculty of Medicine in the Galilee, Bar-Ilan University, Safed, Israel
| | - Ateret Davidovich
- Molecular Virology Lab, Azrieli Faculty of Medicine in the Galilee, Bar-Ilan University, Safed, Israel
| | - Anat Lavi-Itzkovitz
- Molecular Virology Lab, Azrieli Faculty of Medicine in the Galilee, Bar-Ilan University, Safed, Israel
- Bioengineering, Faculty of Engineering, Bar-Ilan University, Ramat-Gan, Israel
| | - Tomer Meirson
- Drug Discovery Laboratory, Azrieli Faculty of Medicine in the Galilee, Bar-Ilan University, Safed, Israel
| | - Jacinta Alison Holmes
- Liver Center, Division of Gastroenterology, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, United States of America
| | - Chia-Yen Dai
- Hepatobiliary Division, Department of Internal Medicine and Hepatitis Center, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
- School of Medicine and Hepatitis Research Center, College of Medicine, and Center for Cancer Research and Center for Liquid Biopsy, Kaohsiung Medical University, Kaohsiung, Taiwan
| | - Chung-Feng Huang
- Hepatobiliary Division, Department of Internal Medicine and Hepatitis Center, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
- School of Medicine and Hepatitis Research Center, College of Medicine, and Center for Cancer Research and Center for Liquid Biopsy, Kaohsiung Medical University, Kaohsiung, Taiwan
| | - Raymond T. Chung
- Liver Center, Division of Gastroenterology, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, United States of America
| | - Assy Nimer
- Internal Medicine Department A, Western Galilee Medical Center, Naharyia, and Azrieli Faculty of Medicine in the Galilee, Bar-Ilan University, Safed, Israel
| | - Assam El-Osta
- Epigenetics in Human Health and Disease Laboratory, Department of Diabetes, Central Clinical School, Monash University, Melbourne, Australia
- Hong Kong Institute of Diabetes and Obesity, Prince of Wales Hospital, The Chinese University of Hong Kong, Hong Kong SAR
| | - Gur Yaari
- Bioengineering, Faculty of Engineering, Bar-Ilan University, Ramat-Gan, Israel
| | - Salomon M. Stemmer
- Davidoff Center, Rabin Medical Center, Beilinson Campus, Petach Tikva, and Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel
| | - Ming-Lung Yu
- Hepatobiliary Division, Department of Internal Medicine and Hepatitis Center, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
- School of Medicine and Hepatitis Research Center, College of Medicine, and Center for Cancer Research and Center for Liquid Biopsy, Kaohsiung Medical University, Kaohsiung, Taiwan
- College of Biological Science and Technology, National Chiao Tung University, Hsin-Chu, Taiwan
- Institute of Biomedical Sciences, National Sun Yat-Sen University, Kaohsiung, Taiwan
| | - Izhak Haviv
- Cancer Personalized Medicine and Diagnostic Genomics Lab, Azrieli Faculty of Medicine in the Galilee, Bar-Ilan University, Safed, Israel
| | - Meital Gal-Tanamy
- Molecular Virology Lab, Azrieli Faculty of Medicine in the Galilee, Bar-Ilan University, Safed, Israel
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21
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Pol S, Lagaye S. The remarkable history of the hepatitis C virus. Genes Immun 2019; 20:436-446. [PMID: 31019253 DOI: 10.1038/s41435-019-0066-z] [Citation(s) in RCA: 20] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/28/2019] [Revised: 03/25/2019] [Accepted: 04/01/2019] [Indexed: 02/06/2023]
Abstract
The infection with the hepatitis C virus (HCV) is an example of the translational research success. The reciprocal interactions between clinicians and scientists have allowed in 30 years the initiation of empirical treatments by interferon, the discovery of the virus, the development of serological and virological tools for diagnosis but also for prognosis (the non-invasive biochemical or morphological fibrosis tests, the predictors of the specific immune response including genetic IL28B polymorphisms). Finally, well-tolerated and effective treatments with oral antivirals inhibiting HCV non-structural viral proteins involved in viral replication have been marketed this last decade, allowing the cure of all infected subjects. HCV chronic infection, which is a public health issue, is a hepatic disease, which may lead to a cirrhosis and an hepatocellular carcinoma (HCC) but also a systemic disease with extra-hepatic manifestations either associated with a cryoglobulinemic vasculitis or chronic inflammation. The HCV infection is the only chronic viral infection, which may be cured: the so-called sustained virologic response, defined by undetectable HCV RNA 12 weeks after the end of the treatment, significantly reduces the risk of morbidity and mortality associated with hepatic and extra-hepatic manifestations, which are mainly reversible. The history of HCV ends with the pangenotypic efficacy of the multiple combinations, easy to use for 8-12 weeks with one to three pills per day and little problems of tolerance. This explains the short 30 years from the virus discovery to the viral hepatitis elimination policy proposed by the World Health Organization (WHO) in 2016.
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Affiliation(s)
- Stanislas Pol
- Université Paris Descartes, Paris, France. .,Département d'Hépatologie, Hôpital Cochin, APHP, Paris, France. .,INSERM UMS-20, Institut Pasteur, Paris, France. .,Immunobiologie des Cellules Dendritiques, Institut Pasteur, Paris, France. .,INSERM U1223, Institut Pasteur, Paris, France.
| | - Sylvie Lagaye
- Immunobiologie des Cellules Dendritiques, Institut Pasteur, Paris, France. .,INSERM U1223, Institut Pasteur, Paris, France.
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22
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Advances in the role of HCV nonstructural protein 5a (NS5A) of 3a genotype in inducing insulin resistance by possible phosphorylation of AKT/PKB. Sci Rep 2019; 9:6150. [PMID: 30992506 PMCID: PMC6468007 DOI: 10.1038/s41598-019-42602-2] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/15/2018] [Accepted: 03/15/2019] [Indexed: 12/15/2022] Open
Abstract
HCV genes interfere with host cellular genes and play crucial role in pathogenesis. The mechanism under which HCV genes induce insulin resistance is not much clear. This study is aimed to examine the role of HCV NS5A in inducing insulin resistance by examining its affect in the phosphorylation level of AKT/PKB. In the present study, HepG2 cells were transfected with HCV NS5A and after 24 hours of transfection, protein was extracted from cells that were pre induced with insulin at three different time intervals i.e. 1hour, 2 hours and 3hours. Dot Blot analysis was performed to study the phosphorylation level of AKT. Results showed that there is clear upregulation of serine 473 phosphorylation level of AKT in NS5A transfected cells as compared with control (without NS5A). In conclusion, upregulation of serine 473 phosphorylation by NS5A of HCV genotype 3a suggests that this gene impairs the normal insulin AKT/PKB signaling pathway that leads towards insulin resistance and Type 2 diabetes mellitus. Therefore, HCV non-structural protein NS5A should be considered as promising candidate to be studied in detail for HCV induced insulin resistance and should be regarded as a therapeutically important target for the prevention of chronic liver diseases.
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23
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Zeisel MB, Dhawan P, Baumert TF. Tight junction proteins in gastrointestinal and liver disease. Gut 2019; 68:547-561. [PMID: 30297438 PMCID: PMC6453741 DOI: 10.1136/gutjnl-2018-316906] [Citation(s) in RCA: 211] [Impact Index Per Article: 35.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/31/2018] [Revised: 08/16/2018] [Accepted: 08/19/2018] [Indexed: 12/11/2022]
Abstract
Over the past two decades a growing body of evidence has demonstrated an important role of tight junction (TJ) proteins in the physiology and disease biology of GI and liver disease. On one side, TJ proteins exert their functional role as integral proteins of TJs in forming barriers in the gut and the liver. Furthermore, TJ proteins can also be expressed outside TJs where they play important functional roles in signalling, trafficking and regulation of gene expression. A hallmark of TJ proteins in disease biology is their functional role in epithelial-to-mesenchymal transition. A causative role of TJ proteins has been established in the pathogenesis of colorectal cancer and gastric cancer. Among the best characterised roles of TJ proteins in liver disease biology is their function as cell entry receptors for HCV-one of the most common causes of hepatocellular carcinoma. At the same time TJ proteins are emerging as targets for novel therapeutic approaches for GI and liver disease. Here we review our current knowledge of the role of TJ proteins in the pathogenesis of GI and liver disease biology and discuss their potential as therapeutic targets.
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Affiliation(s)
- Mirjam B. Zeisel
- Inserm U1052, CNRS UMR 5286, Cancer Research Center of Lyon (CRCL), Université de Lyon (UCBL), Lyon, France
- Inserm, U1110, Institut de Recherche sur les Maladies Virales et Hépatiques, Strasbourg, France
- Université de Strasbourg, Strasbourg, France
| | - Punita Dhawan
- Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, NE
- Buffet Cancer Center, University of Nebraska Medical Center, Omaha, NE
- VA Nebraska-Western Iowa Health Care System, Omaha, NE
| | - Thomas F. Baumert
- Inserm, U1110, Institut de Recherche sur les Maladies Virales et Hépatiques, Strasbourg, France
- Université de Strasbourg, Strasbourg, France
- Institut Hospitalo-Universitaire, Pôle hépato-digestif, Nouvel Hôpital Civil, Strasbourg, France
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24
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The actin cytoskeleton is important for rotavirus internalization and RNA genome replication. Virus Res 2019; 263:27-33. [PMID: 30639190 PMCID: PMC7173133 DOI: 10.1016/j.virusres.2019.01.003] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/10/2018] [Revised: 12/13/2018] [Accepted: 01/08/2019] [Indexed: 02/06/2023]
Abstract
Different stages of the rotavirus lifecycle depend on the dynamics of the actin cytoskeleton. Alpha-actinin, Diaph, and the GTPase Cdc42 are important for virus entry. The GTPAse Rac1 is required for maximal viral RNA synthesis. Numerous host factors are required for the efficient replication of rotavirus, including the activation and inactivation of several cell signaling pathways. One of the cellular structures that are reorganized during rotavirus infection is the actin cytoskeleton. In this work, we report that the dynamics of the actin microfilaments are important at different stages of the virus life cycle, specifically, during virus internalization and viral RNA synthesis at 6 h post-infection. Our results show that the actin-binding proteins alpha-actinin 4 and Diaph, as well as the Rho-family small GTPase Cdc42 are necessary for an efficient virus entry, while GTPase Rac1 is required for maximal viral RNA synthesis.
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25
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Robinson M, Schor S, Barouch-Bentov R, Einav S. Viral journeys on the intracellular highways. Cell Mol Life Sci 2018; 75:3693-3714. [PMID: 30043139 PMCID: PMC6151136 DOI: 10.1007/s00018-018-2882-0] [Citation(s) in RCA: 58] [Impact Index Per Article: 8.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/30/2018] [Revised: 07/01/2018] [Accepted: 07/19/2018] [Indexed: 12/24/2022]
Abstract
Viruses are obligate intracellular pathogens that are dependent on cellular machineries for their replication. Recent technological breakthroughs have facilitated reliable identification of host factors required for viral infections and better characterization of the virus-host interplay. While these studies have revealed cellular machineries that are uniquely required by individual viruses, accumulating data also indicate the presence of broadly required mechanisms. Among these overlapping cellular functions are components of intracellular membrane trafficking pathways. Here, we review recent discoveries focused on how viruses exploit intracellular membrane trafficking pathways to promote various stages of their life cycle, with an emphasis on cellular factors that are usurped by a broad range of viruses. We describe broadly required components of the endocytic and secretory pathways, the Endosomal Sorting Complexes Required for Transport pathway, and the autophagy pathway. Identification of such overlapping host functions offers new opportunities to develop broad-spectrum host-targeted antiviral strategies.
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Affiliation(s)
- Makeda Robinson
- Division of Infectious Diseases and Geographic Medicine, Department of Medicine, Stanford University School of Medicine, 300 Pasteur Drive, Lane Building, Rm L127, Stanford, CA, 94305, USA
- Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, USA
| | - Stanford Schor
- Division of Infectious Diseases and Geographic Medicine, Department of Medicine, Stanford University School of Medicine, 300 Pasteur Drive, Lane Building, Rm L127, Stanford, CA, 94305, USA
| | - Rina Barouch-Bentov
- Division of Infectious Diseases and Geographic Medicine, Department of Medicine, Stanford University School of Medicine, 300 Pasteur Drive, Lane Building, Rm L127, Stanford, CA, 94305, USA
| | - Shirit Einav
- Division of Infectious Diseases and Geographic Medicine, Department of Medicine, Stanford University School of Medicine, 300 Pasteur Drive, Lane Building, Rm L127, Stanford, CA, 94305, USA.
- Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, USA.
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26
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EspH Suppresses Erk by Spatial Segregation from CD81 Tetraspanin Microdomains. Infect Immun 2018; 86:IAI.00303-18. [PMID: 30037792 DOI: 10.1128/iai.00303-18] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/23/2018] [Accepted: 07/10/2018] [Indexed: 01/12/2023] Open
Abstract
Enteropathogenic Escherichia coli (EPEC) belongs to a group of enteric human pathogens known as attaching-and-effacing (A/E) pathogens, which utilize a type III secretion system (T3SS) to translocate a battery of effector proteins from their own cytoplasm into host intestinal epithelial cells. Here we identified EspH to be an effector that prompts the recruitment of the tetraspanin CD81 to infection sites. EspH was also shown to be an effector that suppresses the mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (Erk) signaling pathway at longer infection times. The inhibitory effect was abrogated upon deletion of the last 38 amino acids located at the C terminus of the protein. The efficacy of EspH-dependent Erk suppression was higher in CD81-deficient cells, suggesting that CD81 may act as a positive regulator of Erk, counteracting Erk suppression by EspH. EspH was found within CD81 microdomains soon after infection but was largely excluded from these domains at a later time. Based on our results, we propose a mechanism whereby CD81 is initially recruited to infection sites in response to EspH translocation. At a later stage, EspH moves out of the CD81 clusters to facilitate effective Erk inhibition. Moreover, EspH selectively inhibits the tumor necrosis factor alpha (TNF-α)-induced Erk signaling pathway. Since Erk and TNF-α have been implicated in innate immunity and cell survival, our studies suggest a novel mechanism by which EPEC suppresses these processes to promote its own colonization and survival in the infected gut.
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27
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Rotavirus-Induced Early Activation of the RhoA/ROCK/MLC Signaling Pathway Mediates the Disruption of Tight Junctions in Polarized MDCK Cells. Sci Rep 2018; 8:13931. [PMID: 30224682 PMCID: PMC6141481 DOI: 10.1038/s41598-018-32352-y] [Citation(s) in RCA: 26] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/02/2018] [Accepted: 09/06/2018] [Indexed: 02/02/2023] Open
Abstract
Intestinal epithelial tight junctions (TJ) are a major barrier restricting the entry of various harmful factors including pathogens; however, they also represent an important entry portal for pathogens. Although the rotavirus-induced early disruption of TJ integrity and targeting of TJ proteins as coreceptors are well-defined, the precise molecular mechanisms involved remain unknown. In the present study, infection of polarized MDCK cells with the species A rotavirus (RVA) strains human DS-1 and bovine NCDV induced a redistribution of TJ proteins into the cytoplasm, a reversible decrease in transepithelial resistance, and an increase in paracellular permeability. RhoA/ROCK/MLC signaling was identified as activated at an early stage of infection, while inhibition of this pathway prevented the rotavirus-induced early disruption of TJ integrity and alteration of TJ protein distribution. Activation of pMYPT, PKC, or MLCK, which are known to participate in TJ dissociation, was not observed in MDCK cells infected with either rotavirus strain. Our data demonstrated that binding of RVA virions or cogent VP8* proteins to cellular receptors activates RhoA/ROCK/MLC signaling, which alters TJ protein distribution and disrupts TJ integrity via contraction of the perijunctional actomyosin ring, facilitating virion access to coreceptors and entry into cells.
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28
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Hepatitis C virus enters liver cells using the CD81 receptor complex proteins calpain-5 and CBLB. PLoS Pathog 2018; 14:e1007111. [PMID: 30024968 PMCID: PMC6053247 DOI: 10.1371/journal.ppat.1007111] [Citation(s) in RCA: 41] [Impact Index Per Article: 5.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/11/2017] [Accepted: 05/18/2018] [Indexed: 12/24/2022] Open
Abstract
Hepatitis C virus (HCV) and the malaria parasite Plasmodium use the membrane protein CD81 to invade human liver cells. Here we mapped 33 host protein interactions of CD81 in primary human liver and hepatoma cells using high-resolution quantitative proteomics. In the CD81 protein network, we identified five proteins which are HCV entry factors or facilitators including epidermal growth factor receptor (EGFR). Notably, we discovered calpain-5 (CAPN5) and the ubiquitin ligase Casitas B-lineage lymphoma proto-oncogene B (CBLB) to form a complex with CD81 and support HCV entry. CAPN5 and CBLB were required for a post-binding and pre-replication step in the HCV life cycle. Knockout of CAPN5 and CBLB reduced susceptibility to all tested HCV genotypes, but not to other enveloped viruses such as vesicular stomatitis virus and human coronavirus. Furthermore, Plasmodium sporozoites relied on a distinct set of CD81 interaction partners for liver cell entry. Our findings reveal a comprehensive CD81 network in human liver cells and show that HCV and Plasmodium highjack selective CD81 interactions, including CAPN5 and CBLB for HCV, to invade cells. CD81 is a cell membrane protein, which functions as entry factor for hepatitis C virus (HCV) and malaria sporozoites in the human liver. Currently, it remains enigmatic how CD81 guides the entry process of both pathogens and whether it functions in a similar way during liver cell invasion of HCV and malaria parasites. Here, we use high resolution quantitative proteomics to identify CD81 associated host proteins in liver cells. We found that at least 33 proteins form a complex with CD81, 23 of which were not reported as interaction partners before. We further determined that at least five CD81 interactors are HCV host factors, among them calpain-5 (CAPN5) and the ubiquitin ligase Casitas B-lineage lymphoma proto-oncogene B (CBLB). All tested HCV genotypes require CAPN5 and CBLB for full infection, but neither malaria parasites nor other tested enveloped virus rely on CAPN5 or CBLB. Our study maps the liver cell interactome of CD81 and provides new insight into the distinct cell invasion mechanisms of HCV and malaria parasites.
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29
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Identification of Piperazinylbenzenesulfonamides as New Inhibitors of Claudin-1 Trafficking and Hepatitis C Virus Entry. J Virol 2018; 92:JVI.01982-17. [PMID: 29491159 DOI: 10.1128/jvi.01982-17] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/15/2017] [Accepted: 02/20/2018] [Indexed: 12/13/2022] Open
Abstract
Hepatitis C virus (HCV) infection causes 500,000 deaths annually, in association with end-stage liver diseases. Investigations of the HCV life cycle have widened the knowledge of virology, and here we discovered that two piperazinylbenzenesulfonamides inhibit HCV entry into liver cells. The entry of HCV into host cells is a complex process that is not fully understood but is characterized by multiple spatially and temporally regulated steps involving several known host factors. Through a high-content virus infection screening analysis with a library of 1,120 biologically active chemical compounds, we identified SB258585, an antagonist of serotonin receptor 6 (5-HT6), as a new inhibitor of HCV entry in liver-derived cell lines as well as primary hepatocytes. A functional characterization suggested a role for this compound and the compound SB399885, which share similar structures, as inhibitors of a late HCV entry step, modulating the localization of the coreceptor tight junction protein claudin-1 (CLDN1) in a 5-HT6-independent manner. Both chemical compounds induced an intracellular accumulation of CLDN1, reflecting export impairment. This regulation correlated with the modulation of protein kinase A (PKA) activity. The PKA inhibitor H89 fully reproduced these phenotypes. Furthermore, PKA activation resulted in increased CLDN1 accumulation at the cell surface. Interestingly, an increase of CLDN1 recycling did not correlate with an increased interaction with CD81 or HCV entry. These findings reinforce the hypothesis of a common pathway, shared by several viruses, which involves G-protein-coupled receptor-dependent signaling in late steps of viral entry.IMPORTANCE The HCV entry process is highly complex, and important details of this structured event are poorly understood. By screening a library of biologically active chemical compounds, we identified two piperazinylbenzenesulfonamides as inhibitors of HCV entry. The mechanism of inhibition was not through the previously described activity of these inhibitors as antagonists of serotonin receptor 6 but instead through modulation of PKA activity in a 5-HT6-independent manner, as proven by the lack of 5-HT6 in the liver. We thus highlighted the involvement of the PKA pathway in modulating HCV entry at a postbinding step and in the recycling of the tight junction protein claudin-1 (CLDN1) toward the cell surface. Our work underscores once more the complexity of HCV entry steps and suggests a role for the PKA pathway as a regulator of CLDN1 recycling, with impacts on both cell biology and virology.
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30
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CD81 Receptor Regions outside the Large Extracellular Loop Determine Hepatitis C Virus Entry into Hepatoma Cells. Viruses 2018; 10:v10040207. [PMID: 29677132 PMCID: PMC5923501 DOI: 10.3390/v10040207] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/18/2018] [Revised: 04/14/2018] [Accepted: 04/19/2018] [Indexed: 02/07/2023] Open
Abstract
Hepatitis C virus (HCV) enters human hepatocytes using four essential entry factors, one of which is human CD81 (hCD81). The tetraspanin hCD81 contains a large extracellular loop (LEL), which interacts with the E2 glycoprotein of HCV. The role of the non-LEL regions of hCD81 (intracellular tails, four transmembrane domains, small extracellular loop and intracellular loop) is poorly understood. Here, we studied the contribution of these domains to HCV susceptibility of hepatoma cells by generating chimeras of related tetraspanins with the hCD81 LEL. Our results show that non-LEL regions in addition to the LEL determine susceptibility of cells to HCV. While closely related tetraspanins (X. tropicalis CD81 and D. rerio CD81) functionally complement hCD81 non-LEL regions, distantly related tetraspanins (C. elegans TSP9 amd D. melanogaster TSP96F) do not and tetraspanins with intermediate homology (hCD9) show an intermediate phenotype. Tetraspanin homology and susceptibility to HCV correlate positively. For some chimeras, infectivity correlates with surface expression. In contrast, the hCD9 chimera is fully surface expressed, binds HCV E2 glycoprotein but is impaired in HCV receptor function. We demonstrate that a cholesterol-coordinating glutamate residue in CD81, which hCD9 lacks, promotes HCV infection. This work highlights the hCD81 non-LEL regions as additional HCV susceptibility-determining factors.
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31
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Riad SE, Elhelw DS, Shawer H, El-Ekiaby N, Salah A, Zekri A, Esmat G, Amleh A, Abdelaziz AI. Disruption of Claudin-1 Expression by miRNA-182 Alters the Susceptibility to Viral Infectivity in HCV Cell Models. Front Genet 2018; 9:93. [PMID: 29616082 PMCID: PMC5869927 DOI: 10.3389/fgene.2018.00093] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/16/2017] [Accepted: 03/05/2018] [Indexed: 01/01/2023] Open
Abstract
HCV entry involves a complex interplay between viral and host molecules. During post-binding interactions, the viral E2 complexes with CD81 receptor for delivery to the tight junction proteins CLDN1 and OCLN, which aid in viral internalization. Targeting HCV entry receptors represents an appealing approach to inhibit viral infectivity. This study aimed at investigating the impact of targeting CLDN1 by microRNAs on HCV infectivity. miR-155 was previously shown to target the 3′UTR of CLDN1 mRNA. Therefore, miR-155 was used as a control in this study. In-silico analysis and luciferase reporter assay were utilized to identify potential targeting miRNAs. The impact of the identified miRNAs on CLDN1 mRNA and protein expression was examined by qRT-PCR, indirect immunofluorescence and western blotting, respectively. The role of the selected miRNAs on HCV infectivity was assessed by measuring the viral load following the ectopic expression of the selected miRNAs. miR-182 was identified in-silico and by experimental validation to target CLDN1. Both miR-155 and miR-182 inhibited CLDN1 mRNA and protein expression in infected Huh7 cells. Ectopic expression of miR-155 increased, while miR-182 reduced the viral load. In conclusion, despite repressing CLDN1, the impact of miR-155 and miR-182 on HCV infectivity is contradictory. Ectopic miR-182 expression is suggested as an upstream regulator of the entry factor CLDN1, harnessing HCV infection.
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Affiliation(s)
- Sarah E Riad
- Pharmacology and Toxicology Department, German University in Cairo, New Cairo, Egypt
| | - Dalia S Elhelw
- Pharmaceutical Chemistry Department, German University in Cairo, New Cairo, Egypt
| | - Heba Shawer
- Biology Department, School of Science and Engineering, American University in Cairo, New Cairo, Egypt
| | - Nada El-Ekiaby
- Pharmacology and Toxicology Department, German University in Cairo, New Cairo, Egypt.,School of Medicine, NewGiza University, Cairo, Egypt
| | - Ayman Salah
- Department of Surgery, Cairo University, Cairo, Egypt
| | - Abdelrahman Zekri
- Virology and Immunology Unit, Cancer Biology Department, National Cancer Institute, Cairo University, Cairo, Egypt
| | - Gamal Esmat
- Department of Endemic Medicine and Hepatology, Cairo University, Cairo, Egypt
| | - Asma Amleh
- Biology Department, School of Science and Engineering, American University in Cairo, New Cairo, Egypt
| | - Ahmed I Abdelaziz
- Pharmacology and Toxicology Department, German University in Cairo, New Cairo, Egypt.,School of Medicine, NewGiza University, Cairo, Egypt
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32
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Morozov VA, Lagaye S. Hepatitis C virus: Morphogenesis, infection and therapy. World J Hepatol 2018; 10:186-212. [PMID: 29527256 PMCID: PMC5838439 DOI: 10.4254/wjh.v10.i2.186] [Citation(s) in RCA: 87] [Impact Index Per Article: 12.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/04/2017] [Revised: 01/11/2018] [Accepted: 02/07/2018] [Indexed: 02/06/2023] Open
Abstract
Hepatitis C virus (HCV) is a major cause of liver diseases including liver cirrhosis and hepatocellular carcinoma. Approximately 3% of the world population is infected with HCV. Thus, HCV infection is considered a public healthy challenge. It is worth mentioning, that the HCV prevalence is dependent on the countries with infection rates around 20% in high endemic countries. The review summarizes recent data on HCV molecular biology, the physiopathology of infection (immune-mediated liver damage, liver fibrosis and lipid metabolism), virus diagnostic and treatment. In addition, currently available in vitro, ex vivo and animal models to study the virus life cycle, virus pathogenesis and therapy are described. Understanding of both host and viral factors may in the future lead to creation of new approaches in generation of an efficient therapeutic vaccine.
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Affiliation(s)
- Vladimir Alexei Morozov
- Center for HIV and Retrovirology, Department of Infectious Diseases, Robert Koch Institute, Berlin 13353, Germany
| | - Sylvie Lagaye
- Department of Immunology, Institut Pasteur, INSERM U1223, Paris 75015, France
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33
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Min S, Lim YS, Shin D, Park C, Park JB, Kim S, Windisch MP, Hwang SB. Abl Tyrosine Kinase Regulates Hepatitis C Virus Entry. Front Microbiol 2017; 8:1129. [PMID: 28674529 PMCID: PMC5474468 DOI: 10.3389/fmicb.2017.01129] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/10/2017] [Accepted: 06/02/2017] [Indexed: 12/12/2022] Open
Abstract
Abl is a central regulator of multiple cellular processes controlling actin dynamics, proliferation, and differentiation. Here, we showed that knockdown of Abl impaired hepatitis C virus (HCV) propagation. Treatment of Abl tyrosine kinase-specific inhibitor, imatinib and dasatinib, also significantly decreased HCV RNA and protein levels in HCV-infected cells. We showed that both imatinib and dasatinib selectively inhibited HCV infection at the entry step of HCV life cycle, suggesting that Abl kinase activity may be necessary for HCV entry. Using HCV pseudoparticle infection assays, we verified that Abl is required for viral entry. By employing transferrin uptake and immunofluorescence assays, we further demonstrated that Abl was involved in HCV entry at a clathrin-mediated endocytosis step. These data suggest that Abl may represent a novel host factor for HCV entry.
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Affiliation(s)
- Saehong Min
- Department of Biomedical Gerontology, Graduate School of Hallym UniversityChuncheon, South Korea.,National Research Laboratory of Hepatitis C Virus and Ilsong Institute of Life Science, Hallym UniversityAnyang, South Korea
| | - Yun-Sook Lim
- National Research Laboratory of Hepatitis C Virus and Ilsong Institute of Life Science, Hallym UniversityAnyang, South Korea
| | - Dongjo Shin
- Department of Biomedical Gerontology, Graduate School of Hallym UniversityChuncheon, South Korea.,Hepatitis Research Laboratory, Institut Pasteur KoreaSeongnam, South Korea
| | - Chorong Park
- Department of Biomedical Gerontology, Graduate School of Hallym UniversityChuncheon, South Korea.,National Research Laboratory of Hepatitis C Virus and Ilsong Institute of Life Science, Hallym UniversityAnyang, South Korea
| | - Jae-Bong Park
- Department of Biochemistry, College of Medicine, Hallym UniversityChuncheon, South Korea
| | - Seungtaek Kim
- Institute of Gastroenterology, Yonsei University College of MedicineSeoul, South Korea
| | - Marc P Windisch
- Hepatitis Research Laboratory, Institut Pasteur KoreaSeongnam, South Korea
| | - Soon B Hwang
- Department of Biomedical Gerontology, Graduate School of Hallym UniversityChuncheon, South Korea.,National Research Laboratory of Hepatitis C Virus and Ilsong Institute of Life Science, Hallym UniversityAnyang, South Korea
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34
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Regulated Entry of Hepatitis C Virus into Hepatocytes. Viruses 2017; 9:v9050100. [PMID: 28486435 PMCID: PMC5454413 DOI: 10.3390/v9050100] [Citation(s) in RCA: 32] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/21/2017] [Revised: 04/24/2017] [Accepted: 05/02/2017] [Indexed: 02/07/2023] Open
Abstract
Hepatitis C virus (HCV) is a model for the study of virus–host interaction and host cell responses to infection. Virus entry into hepatocytes is the first step in the HCV life cycle, and this process requires multiple receptors working together. The scavenger receptor class B type I (SR-BI) and claudin-1 (CLDN1), together with human cluster of differentiation (CD) 81 and occludin (OCLN), constitute the minimal set of HCV entry receptors. Nevertheless, HCV entry is a complex process involving multiple host signaling pathways that form a systematic regulatory network; this network is centrally controlled by upstream regulators epidermal growth factor receptor (EGFR) and transforming growth factor β receptor (TGFβ-R). Further feedback regulation and cell-to-cell spread of the virus contribute to the chronic maintenance of HCV infection. A comprehensive and accurate disclosure of this critical process should provide insights into the viral entry mechanism, and offer new strategies for treatment regimens and targets for HCV therapeutics.
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35
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Denolly S, Cosset FL. A master regulator of tight junctions involved in hepatitis C virus entry and pathogenesis. Hepatology 2017; 65:1756-1758. [PMID: 28177536 DOI: 10.1002/hep.29066] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/28/2016] [Revised: 01/07/2017] [Accepted: 01/11/2017] [Indexed: 12/26/2022]
Affiliation(s)
- Solène Denolly
- CIRI-International Center for Infectiology Research, Team EVIR, Inserm, U1111, Université Claude Bernard Lyon-1, CNRS UMR5308, Ecole Normale Supérieure de Lyon, Université de Lyon, Lyon, France
| | - François-Loïc Cosset
- CIRI-International Center for Infectiology Research, Team EVIR, Inserm, U1111, Université Claude Bernard Lyon-1, CNRS UMR5308, Ecole Normale Supérieure de Lyon, Université de Lyon, Lyon, France
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36
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Chiang AWT, Wu WYL, Wang T, Hwang MJ. Identification of Entry Factors Involved in Hepatitis C Virus Infection Based on Host-Mimicking Short Linear Motifs. PLoS Comput Biol 2017; 13:e1005368. [PMID: 28129350 PMCID: PMC5302801 DOI: 10.1371/journal.pcbi.1005368] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/08/2016] [Revised: 02/10/2017] [Accepted: 01/17/2017] [Indexed: 12/15/2022] Open
Abstract
Host factors that facilitate viral entry into cells can, in principle, be identified from a virus-host protein interaction network, but for most viruses information for such a network is limited. To help fill this void, we developed a bioinformatics approach and applied it to hepatitis C virus (HCV) infection, which is a current concern for global health. Using this approach, we identified short linear sequence motifs, conserved in the envelope proteins of HCV (E1/E2), that potentially can bind human proteins present on the surface of hepatocytes so as to construct an HCV (envelope)-host protein interaction network. Gene Ontology functional and KEGG pathway analyses showed that the identified host proteins are enriched in cell entry and carcinogenesis functionalities. The validity of our results is supported by much published experimental data. Our general approach should be useful when developing antiviral agents, particularly those that target virus-host interactions. Viruses recruit host proteins, called entry factors, to help gain entry to host cells. Identification of entry factors can provide targets for developing antiviral drugs. By exploring the concept that short linear peptide motifs involved in human protein-protein interactions may be mimicked by viruses to hijack certain host cellular processes and thereby assist viral infection/survival, we developed a bioinformatics strategy to computationally identify entry factors of hepatitis C virus (HCV) infection, which is a worldwide health problem. Analysis of cellular functions and biochemical pathways indicated that the human proteins we identified usually play a role in cell entry and/or carcinogenesis, and results of the analysis are generally supported by experimental studies on HCV infection, including the ~80% (15 of 19) prediction rate of known HCV hepatocyte entry factors. Because molecular mimicry is a general concept, our bioinformatics strategy is a timely approach to identify new targets for antiviral research, not only for HCV but also for other viruses.
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Affiliation(s)
| | - Walt Y. L. Wu
- Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan
| | - Ting Wang
- Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan
| | - Ming-Jing Hwang
- Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan
- * E-mail:
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37
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Xu Q, Cao M, Song H, Chen S, Qian X, Zhao P, Ren H, Tang H, Wang Y, Wei Y, Zhu Y, Qi Z. Caveolin-1-mediated Japanese encephalitis virus entry requires a two-step regulation of actin reorganization. Future Microbiol 2016; 11:1227-1248. [DOI: 10.2217/fmb-2016-0002] [Citation(s) in RCA: 33] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022] Open
Abstract
Aim: To investigate the detailed mechanism of Japanese encephalitis virus (JEV) cell entry. Materials & methods: Utilize a siRNA library targeting cellular membrane trafficking genes to identify key molecules that mediate JEV entry into human neuronal cells. Results: JEV enters human neuronal cells by caveolin-1-mediated endocytosis, which depends on a two-step regulation of actin cytoskeleton remodeling triggered by RhoA and Rac1: RhoA activation promoted the phosphorylation of caveolin-1, and then Rac1 activation facilitated caveolin-associated viral internalization. Specifically, virus attachment activates the EGFR–PI3K signaling pathway, thereby leading to RhoA activation. Conclusion: This work provides a detailed picture of the entry route and intricate cellular events following the entry of JEV into human neuronal cells, and promotes a better understanding of JEV entry.
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Affiliation(s)
- Qingqiang Xu
- Department of Microbiology, Second Military Medical University, Shanghai Key Laboratory of Medical Biodefense, Shanghai 200433, China
| | - Mingmei Cao
- Department of Medical Microbiology & Parasitology, Second Military Medical University, Shanghai 200433, China
| | - Hongyuan Song
- Department of Ophthalmology, Changhai Hospital, Second Military Medical University, Shanghai 200433, China
| | - Shenglin Chen
- Department of Microbiology, Second Military Medical University, Shanghai Key Laboratory of Medical Biodefense, Shanghai 200433, China
| | - Xijing Qian
- Department of Microbiology, Second Military Medical University, Shanghai Key Laboratory of Medical Biodefense, Shanghai 200433, China
| | - Ping Zhao
- Department of Microbiology, Second Military Medical University, Shanghai Key Laboratory of Medical Biodefense, Shanghai 200433, China
| | - Hao Ren
- Department of Microbiology, Second Military Medical University, Shanghai Key Laboratory of Medical Biodefense, Shanghai 200433, China
| | - Hailin Tang
- Department of Microbiology, Second Military Medical University, Shanghai Key Laboratory of Medical Biodefense, Shanghai 200433, China
| | - Yan Wang
- Department of Microbiology, Second Military Medical University, Shanghai Key Laboratory of Medical Biodefense, Shanghai 200433, China
| | - Youheng Wei
- State Key Laboratory of Genetic Engineering, Institute of Genetics, School of Life Sciences, Fudan University, Shanghai 200438, China
| | - Yongzhe Zhu
- Department of Microbiology, Second Military Medical University, Shanghai Key Laboratory of Medical Biodefense, Shanghai 200433, China
| | - Zhongtian Qi
- Department of Microbiology, Second Military Medical University, Shanghai Key Laboratory of Medical Biodefense, Shanghai 200433, China
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38
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Sugiyama M, Kanto T. Dual blockade of hepatitis C virus entry at a gatekeeper of hepatocytes: Not only a preventive, but also therapeutic target of claudin 1. Hepatology 2016; 64:979-82. [PMID: 27405253 DOI: 10.1002/hep.28726] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/07/2022]
Affiliation(s)
- Masaya Sugiyama
- Genome Medical Science Project, The Research Center for Hepatitis and Immunology, National Center for Global Health and Medicine, Ichikawa, Japan.,Department of Liver Diseases, The Research Center for Hepatitis and Immunology, National Center for Global Health and Medicine, Ichikawa, Japan
| | - Tatsuya Kanto
- Department of Liver Diseases, The Research Center for Hepatitis and Immunology, National Center for Global Health and Medicine, Ichikawa, Japan
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39
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Hepatitis C virus cell entry: a target for novel antiviral strategies to address limitations of direct acting antivirals. Hepatol Int 2016; 10:741-8. [DOI: 10.1007/s12072-016-9724-7] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/15/2015] [Accepted: 03/16/2016] [Indexed: 12/12/2022]
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40
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Grassi G, Di Caprio G, Fimia GM, Ippolito G, Tripodi M, Alonzi T. Hepatitis C virus relies on lipoproteins for its life cycle. World J Gastroenterol 2016; 22:1953-1965. [PMID: 26877603 PMCID: PMC4726671 DOI: 10.3748/wjg.v22.i6.1953] [Citation(s) in RCA: 36] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/25/2015] [Revised: 10/19/2015] [Accepted: 12/21/2015] [Indexed: 02/06/2023] Open
Abstract
Hepatitis C virus (HCV) infects over 150 million people worldwide. In most cases, HCV infection becomes chronic causing liver disease ranging from fibrosis to cirrhosis and hepatocellular carcinoma. Viral persistence and pathogenesis are due to the ability of HCV to deregulate specific host processes, mainly lipid metabolism and innate immunity. In particular, HCV exploits the lipoprotein machineries for almost all steps of its life cycle. The aim of this review is to summarize current knowledge concerning the interplay between HCV and lipoprotein metabolism. We discuss the role played by members of lipoproteins in HCV entry, replication and virion production.
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Park GB, Kim D, Park SJ, Lee HK, Kim JH, Kim YS, Park SG, Choi IH, Yoon SH, Lee YJ, Paeng S, Hur DY. Pre-stimulation of CD81 expression by resting B cells increases proliferation following EBV infection, but the overexpression of CD81 induces the apoptosis of EBV-transformed B cells. Int J Mol Med 2015; 36:1464-78. [PMID: 26498453 PMCID: PMC4678167 DOI: 10.3892/ijmm.2015.2372] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/11/2015] [Accepted: 09/23/2015] [Indexed: 02/06/2023] Open
Abstract
Hepatitis C virus (HCV) E2 protein binds to CD81, which is a component of the B cell co-stimulatory complex. The E2-CD81 interaction leads to B cell proliferation, protein tyrosine phosphorylation and to the hypermutation of immunoglobulin genes. Epidemiological studies have reported a high prevalence of B cell non-Hodgkin lymphoma (NHL) in HCV-positive patients, suggesting a potential association between HCV and Epstein-Barr virus (EBV) in the genesis of B lymphocyte proliferative disorders. In the present study, in order to investigate the association between EBV and HCV in B cells, we created an in vitro EBV-induced B cell transformation model. CD81 was gradually overexpressed during transformation by EBV. B cells isolated from HCV-positive patients grew more rapidly and clumped together earlier than B cells isolated from healthy donors following EBV infection. Pre-stimulation of CD81 expressed by resting B cells with anti-CD81 monoclonal antibody (mAb) or HCV E2 accelerated the generation of lymphoblastoid cell lines (LCLs) by EBV infection. These cells proliferated prominently through the early expression of interleukin-10 and intracellular latent membrane protein (LMP)-l. By contrast, the overexpression of CD81 on EBV-transformed B cells by anti-CD81 mAb or HCV E2 protein induced apoptosis through reactive oxygen species (ROS)-mediated mitochondrial dysfunction. These results suggest that the engagement of CD81 expressed by B cells has differential effects on B cell fate (proliferation or apoptosis) according to EBV infection and the expression level of CD81.
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Affiliation(s)
- Ga Bin Park
- Department of Anatomy and Research Center for Tumor Immunology, Inje University College of Medicine, Busan 614-735, Republic of Korea
| | - Daejin Kim
- Department of Anatomy and Research Center for Tumor Immunology, Inje University College of Medicine, Busan 614-735, Republic of Korea
| | - Sung Jae Park
- Department of Internal Medicine, Inje University Busan Paik Hospital, Busan 614-735, Republic of Korea
| | - Hyun-Kyung Lee
- Department of Internal Medicine, Inje University Busan Paik Hospital, Busan 614-735, Republic of Korea
| | - Ji Hyun Kim
- Department of Internal Medicine, Inje University Busan Paik Hospital, Busan 614-735, Republic of Korea
| | - Yeong Seok Kim
- Department of Anatomy and Research Center for Tumor Immunology, Inje University College of Medicine, Busan 614-735, Republic of Korea
| | - Sae-Gwang Park
- Department of Microbiology, Inje University College of Medicine, Busan 614-735, Republic of Korea
| | - In-Hak Choi
- Department of Microbiology, Inje University College of Medicine, Busan 614-735, Republic of Korea
| | - Sung Ho Yoon
- Department of Plastic Surgery, Inje University Haeundae Paik Hospital, Busan 614-735, Republic of Korea
| | - Youn Jae Lee
- Department of Internal Medicine, Inje University Busan Paik Hospital, Busan 614-735, Republic of Korea
| | - Sunghwa Paeng
- Department of Neurosurgery, Inje University Busan Paik Hospital, Busan 614-735, Republic of Korea
| | - Dae Young Hur
- Department of Anatomy and Research Center for Tumor Immunology, Inje University College of Medicine, Busan 614-735, Republic of Korea
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42
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Colpitts CC, Verrier ER, Baumert TF. Targeting Viral Entry for Treatment of Hepatitis B and C Virus Infections. ACS Infect Dis 2015; 1:420-7. [PMID: 27617925 DOI: 10.1021/acsinfecdis.5b00039] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
Hepatitis B virus (HBV) and hepatitis C virus (HCV) infections remain major health problems worldwide, with 400-500 million chronically infected people worldwide. Chronic infection results in liver cirrhosis and hepatocellular carcinoma, the second leading cause of cancer death. Current treatments for HBV limit viral replication without efficiently curing infection. HCV treatment has markedly progressed with the licensing of direct-acting antivirals (DAAs) for HCV cure, yet limited access for the majority of patients is a major challenge. Preventative and curative treatment strategies, aimed at novel targets, are needed for both viruses. Viral entry represents one such target, although detailed knowledge of the entry mechanisms is a prerequisite. For HBV, the recent discovery of the NTCP cell entry factor enabled the establishment of an HBV cell culture model and showed that cyclosporin A and Myrcludex B are NTCP-targeting entry inhibitors. Advances in the understanding of HCV entry revealed it to be a complex process involving many factors, offering several antiviral targets. These include viral envelope proteins E1 and E2, virion-associated lipoprotein ApoE, and cellular factors CD81, SRBI, EGFR, claudin-1, occludin, and the cholesterol transporter NPC1L1. Small molecules targeting SR-BI, EGFR, and NPC1L1 have entered clinical trials, whereas other viral- and host-targeted small molecules, peptides, and antibodies show promise in preclinical models. This review summarizes the current understanding of HBV and HCV entry and describes novel antiviral targets and compounds in different stages of clinical development. Overall, proof-of-concept studies indicate that entry inhibitors are a promising class of antivirals to prevent and treat HBV and HCV infections.
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Affiliation(s)
- Che C. Colpitts
- Inserm, U1110, Institut de Recherche sur les Maladies
Virales et Hépatiques, 67000 Strasbourg, France
- Université de Strasbourg, 67000 Strasbourg, France
| | - Eloi R. Verrier
- Inserm, U1110, Institut de Recherche sur les Maladies
Virales et Hépatiques, 67000 Strasbourg, France
- Université de Strasbourg, 67000 Strasbourg, France
| | - Thomas F. Baumert
- Inserm, U1110, Institut de Recherche sur les Maladies
Virales et Hépatiques, 67000 Strasbourg, France
- Université de Strasbourg, 67000 Strasbourg, France
- Institut Hospitalo-Universitaire,
Pôle Hépato-digestif, Hopitaux Universitaires de Strasbourg, 67000 Strasbourg, France
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43
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Decoding protein networks during virus entry by quantitative proteomics. Virus Res 2015; 218:25-39. [PMID: 26365680 PMCID: PMC4914609 DOI: 10.1016/j.virusres.2015.09.006] [Citation(s) in RCA: 21] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/14/2015] [Revised: 08/31/2015] [Accepted: 09/08/2015] [Indexed: 01/05/2023]
Abstract
Virus entry into host cells relies on interactions between viral and host structures including lipids, carbohydrates and proteins. Particularly, protein–protein interactions between viral surface proteins and host proteins as well as secondary host protein–protein interactions play a pivotal role in coordinating virus binding and uptake. These interactions are dynamic and frequently involve multiprotein complexes. In the past decade mass spectrometry based proteomics methods have reached sensitivities and high throughput compatibilities of genomics methods and now allow the reliable quantitation of proteins in complex samples from limited material. As proteomics provides essential information on the biologically active entity namely the protein, including its posttranslational modifications and its interactions with other proteins, it is an indispensable method in the virologist's toolbox. Here we review protein interactions during virus entry and compare classical biochemical methods to study entry with novel technically advanced quantitative proteomics techniques. We highlight the value of quantitative proteomics in mapping functional virus entry networks, discuss the benefits and limitations and illustrate how the methodology will help resolve unsettled questions in virus entry research in the future.
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44
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Gerold G, Meissner F, Bruening J, Welsch K, Perin PM, Baumert TF, Vondran FW, Kaderali L, Marcotrigiano J, Khan AG, Mann M, Rice CM, Pietschmann T. Quantitative Proteomics Identifies Serum Response Factor Binding Protein 1 as a Host Factor for Hepatitis C Virus Entry. Cell Rep 2015. [PMID: 26212323 PMCID: PMC4836839 DOI: 10.1016/j.celrep.2015.06.063] [Citation(s) in RCA: 47] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
Hepatitis C virus (HCV) enters human hepatocytes through a multistep mechanism involving, among other host proteins, the virus receptor CD81. How CD81 governs HCV entry is poorly characterized, and CD81 protein interactions after virus binding remain elusive. We have developed a quantitative proteomics protocol to identify HCV-triggered CD81 interactions and found 26 dynamic binding partners. At least six of these proteins promote HCV infection, as indicated by RNAi. We further characterized serum response factor binding protein 1 (SRFBP1), which is recruited to CD81 during HCV uptake and supports HCV infection in hepatoma cells and primary human hepatocytes. SRFBP1 facilitates host cell penetration by all seven HCV genotypes, but not of vesicular stomatitis virus and human coronavirus. Thus, SRFBP1 is an HCV-specific, pan-genotypic host entry factor. These results demonstrate the use of quantitative proteomics to elucidate pathogen entry and underscore the importance of host protein-protein interactions during HCV invasion.
Hepatitis C virus binding alters host protein interactions with the receptor CD81 Six out of 26 virus-dependent CD81-interacting proteins promote virus entry SRFBP1 binds CD81 and aids infection of all HCV, but not VSV, genotypes SRFBP1 is membrane-associated and required for HCV entry
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Affiliation(s)
- Gisa Gerold
- Insitute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, 30165 Hannover, Germany; Center for the Study of Hepatitis C, Laboratory of Virology and Infectious Disease, the Rockefeller University, New York, NY 10065, USA.
| | - Felix Meissner
- Department of Proteomics and Signal Transduction, Max Planck Institute for Biochemistry, 82152 Martinsried, Germany
| | - Janina Bruening
- Insitute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, 30165 Hannover, Germany
| | - Kathrin Welsch
- Insitute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, 30165 Hannover, Germany
| | - Paula M Perin
- Insitute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, 30165 Hannover, Germany
| | - Thomas F Baumert
- Inserm Unit 1110, Université de Strasbourg, Strasbourg 67000, France
| | - Florian W Vondran
- Department of General, Visceral and Transplant Surgery, Hannover Medical School, 30165 Hannover, Germany
| | - Lars Kaderali
- Institute for Medical Informatics and Biometry (IMB), Medical School, University of Technology Dresden, 01307 Dresden, Germany
| | - Joseph Marcotrigiano
- Center for Advanced Biotechnology and Medicine, Department of Chemistry and Chemical Biology, Rutgers University, Piscataway, NJ 08854, USA
| | - Abdul G Khan
- Center for Advanced Biotechnology and Medicine, Department of Chemistry and Chemical Biology, Rutgers University, Piscataway, NJ 08854, USA
| | - Matthias Mann
- Department of Proteomics and Signal Transduction, Max Planck Institute for Biochemistry, 82152 Martinsried, Germany
| | - Charles M Rice
- Center for the Study of Hepatitis C, Laboratory of Virology and Infectious Disease, the Rockefeller University, New York, NY 10065, USA
| | - Thomas Pietschmann
- Insitute for Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a joint venture between the Medical School Hannover and the Helmholtz Centre for Infection Research, 30165 Hannover, Germany.
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45
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Abstract
Viruses have evolved intricate mechanisms to gain entry into the host cell. Identification of host proteins that serve as viral receptors has enabled insights into virus particle internalization, host and tissue tropism, and viral pathogenesis. In this review we discuss the most commonly employed methods for virus receptor discovery, specifically highlighting the use of forward genetic screens in human haploid cells. The ability to generate true knockout alleles at high saturation provides a sensitive means to study virus-host interactions. To illustrate the power of such haploid genetic screens, we highlight the discovery of the lysosomal proteins NPC1 and LAMP1 as intracellular receptors for Ebola virus and Lassa virus, respectively. From these studies emerges the notion that receptor usage by these viruses is highly dynamic, involving a programmed switch from cell surface receptor to intracellular receptor. Broad application of genetic knockout approaches will chart functional landscapes of receptors and endocytic pathways hijacked by viruses.
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Affiliation(s)
- Sirika Pillay
- Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, California 94305; ,
| | - Jan E Carette
- Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, California 94305; ,
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46
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Interaction of human tumor viruses with host cell surface receptors and cell entry. Viruses 2015; 7:2592-617. [PMID: 26008702 PMCID: PMC4452921 DOI: 10.3390/v7052592] [Citation(s) in RCA: 39] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/23/2015] [Accepted: 05/12/2015] [Indexed: 02/06/2023] Open
Abstract
Currently, seven viruses, namely Epstein-Barr virus (EBV), Kaposi's sarcoma-associated herpes virus (KSHV), high-risk human papillomaviruses (HPVs), Merkel cell polyomavirus (MCPyV), hepatitis B virus (HBV), hepatitis C virus (HCV) and human T cell lymphotropic virus type 1 (HTLV-1), have been described to be consistently associated with different types of human cancer. These oncogenic viruses belong to distinct viral families, display diverse cell tropism and cause different malignancies. A key to their pathogenicity is attachment to the host cell and entry in order to replicate and complete their life cycle. Interaction with the host cell during viral entry is characterized by a sequence of events, involving viral envelope and/or capsid molecules as well as cellular entry factors that are critical in target cell recognition, thereby determining cell tropism. Most oncogenic viruses initially attach to cell surface heparan sulfate proteoglycans, followed by conformational change and transfer of the viral particle to secondary high-affinity cell- and virus-specific receptors. This review summarizes the current knowledge of the host cell surface factors and molecular mechanisms underlying oncogenic virus binding and uptake by their cognate host cell(s) with the aim to provide a concise overview of potential target molecules for prevention and/or treatment of oncogenic virus infection.
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47
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Host cell kinases and the hepatitis C virus life cycle. BIOCHIMICA ET BIOPHYSICA ACTA-PROTEINS AND PROTEOMICS 2015; 1854:1657-62. [PMID: 25896387 DOI: 10.1016/j.bbapap.2015.04.011] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 02/07/2015] [Revised: 04/08/2015] [Accepted: 04/09/2015] [Indexed: 02/07/2023]
Abstract
Hepatitis C virus (HCV) infection relies on virus-host interactions with human hepatocytes, a context in which host cell kinases play critical roles in every step of the HCV life cycle. During viral entry, cellular kinases, including EGFR, EphA2 and PKA, regulate the localization of host HCV entry factors and induce receptor complex assembly. Following virion internalization, viral genomes replicate on endoplasmic reticulum-derived membranous webs. The formation of membranous webs depends on interactions between the HCV NS5a protein and PI4KIIIα. The phosphorylation status of NS5a, regulated by PI4KIIIα, CKI and other kinases, also acts as a molecular switch to virion assembly, which takes place on lipid droplets. The formation of lipid droplets is enhanced by HCV activation of IKKα. In view of the multiple crucial steps in the viral life cycle that are mediated by host cell kinases, these enzymes also represent complementary targets for antiviral therapy. This article is part of a Special Issue entitled: Inhibitors of Protein Kinases.
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48
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Jacob T, Broeke CVD, Waesberghe CV, Troys LV, Favoreel HW. Pseudorabies virus US3 triggers RhoA phosphorylation to reorganize the actin cytoskeleton. J Gen Virol 2015; 96:2328-2335. [PMID: 25883194 DOI: 10.1099/vir.0.000152] [Citation(s) in RCA: 21] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
The conserved alphaherpesvirus serine/threonine kinase US3 causes dramatic changes in the actin cytoskeleton, consisting of actin stress fibre breakdown and protrusion formation, associated with increased virus spread. Here, we showed that US3 expression led to RhoA phosphorylation at serine 188 (S188), one of the hallmarks of suppressed RhoA signalling, and that expression of a non-phosphorylatable RhoA variant interfered with the ability of US3 to induce actin rearrangements. Furthermore, inhibition of cellular protein kinase A (PKA) eliminated the ability of US3 to induce S188 RhoA phosphorylation, pointing to a role for PKA in US3-induced RhoA phosphorylation. Hence, the US3 kinase leads to PKA-dependent S188 RhoA phosphorylation, which contributes to US3-mediated actin rearrangements. Our data suggest that US3 efficiently usurps the antagonistic RhoA and Cdc42/Rac1/p21-activated kinase signalling branches to rearrange the actin cytoskeleton.
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Affiliation(s)
- Thary Jacob
- Department of Virology, Parasitology, and Immunology, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, 9820 Merelbeke, Belgium
| | - Céline Van den Broeke
- Department of Virology, Parasitology, and Immunology, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, 9820 Merelbeke, Belgium
| | - Cliff Van Waesberghe
- Department of Virology, Parasitology, and Immunology, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, 9820 Merelbeke, Belgium
| | - Leen Van Troys
- Department of Biochemistry, Faculty of Medicine and Health Sciences, Ghent University, Albert Baertsoenkaai 3, 9000 Ghent, Belgium
| | - Herman W Favoreel
- Department of Virology, Parasitology, and Immunology, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, 9820 Merelbeke, Belgium
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49
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Mailly L, Xiao F, Lupberger J, Wilson GK, Aubert P, Duong FHT, Calabrese D, Leboeuf C, Fofana I, Thumann C, Bandiera S, Lütgehetmann M, Volz T, Davis C, Harris HJ, Mee CJ, Girardi E, Chane-Woon-Ming B, Ericsson M, Fletcher N, Bartenschlager R, Pessaux P, Vercauteren K, Meuleman P, Villa P, Kaderali L, Pfeffer S, Heim MH, Neunlist M, Zeisel MB, Dandri M, McKeating JA, Robinet E, Baumert TF. Clearance of persistent hepatitis C virus infection in humanized mice using a claudin-1-targeting monoclonal antibody. Nat Biotechnol 2015; 33:549-554. [PMID: 25798937 PMCID: PMC4430301 DOI: 10.1038/nbt.3179] [Citation(s) in RCA: 122] [Impact Index Per Article: 12.2] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/01/2014] [Accepted: 12/22/2014] [Indexed: 12/12/2022]
Abstract
Hepatitis C virus (HCV) infection is a leading cause of liver cirrhosis and cancer1. Cell entry of HCV2 and other pathogens3-5 is mediated by tight junction (TJ) proteins, but successful therapeutic targeting of TJ proteins has not been reported yet. Using a human liver-chimeric mouse model6 we show that a monoclonal antibody specific for TJ protein claudin-17 eliminates chronic HCV infection without detectable toxicity. This antibody inhibits HCV entry, cell-cell transmission and virus-induced signaling events. Antibody treatment reduces the number of HCV-infected hepatocytes in vivo, highlighting the need for de novo infection via host entry factors to maintain chronic infection. In summary, we demonstrate that an antibody targeting a virus receptor can cure chronic viral infection and uncover TJ proteins as targets for antiviral therapy.
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Affiliation(s)
- Laurent Mailly
- Institut National de la Santé et de la Recherche Médicale, U1110, Strasbourg, France.,Université de Strasbourg, Strasbourg, France
| | - Fei Xiao
- Institut National de la Santé et de la Recherche Médicale, U1110, Strasbourg, France.,Université de Strasbourg, Strasbourg, France
| | - Joachim Lupberger
- Institut National de la Santé et de la Recherche Médicale, U1110, Strasbourg, France.,Université de Strasbourg, Strasbourg, France
| | - Garrick K Wilson
- Hepatitis C Research Group, Institute for Biomedical Research, University of Birmingham, Birmingham, United Kingdom
| | - Philippe Aubert
- Institut National de la Santé et de la Recherche Médicale, U913, Nantes, France.,Université de Nantes, Nantes, France.,Institut des Maladies de l'Appareil Digestif, CHU Nantes, Hôpital Hôtel-Dieu, Nantes, France
| | - François H T Duong
- Department of Biomedicine, Hepatology Laboratory, University of Basel, Basel, Switzerland
| | - Diego Calabrese
- Department of Biomedicine, Hepatology Laboratory, University of Basel, Basel, Switzerland
| | - Céline Leboeuf
- Institut National de la Santé et de la Recherche Médicale, U1110, Strasbourg, France.,Université de Strasbourg, Strasbourg, France
| | - Isabel Fofana
- Institut National de la Santé et de la Recherche Médicale, U1110, Strasbourg, France.,Université de Strasbourg, Strasbourg, France
| | - Christine Thumann
- Institut National de la Santé et de la Recherche Médicale, U1110, Strasbourg, France.,Université de Strasbourg, Strasbourg, France
| | - Simonetta Bandiera
- Institut National de la Santé et de la Recherche Médicale, U1110, Strasbourg, France.,Université de Strasbourg, Strasbourg, France
| | - Marc Lütgehetmann
- I. Department of Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Tassilo Volz
- I. Department of Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Christopher Davis
- Hepatitis C Research Group, Institute for Biomedical Research, University of Birmingham, Birmingham, United Kingdom
| | - Helen J Harris
- Hepatitis C Research Group, Institute for Biomedical Research, University of Birmingham, Birmingham, United Kingdom
| | - Christopher J Mee
- Hepatitis C Research Group, Institute for Biomedical Research, University of Birmingham, Birmingham, United Kingdom
| | - Erika Girardi
- Université de Strasbourg, Strasbourg, France.,Architecture et Réactivité de l'ARN, Institut de Biologie Moléculaire et Cellulaire du CNRS - UPR 9002, Strasbourg, France
| | - Béatrice Chane-Woon-Ming
- Université de Strasbourg, Strasbourg, France.,Architecture et Réactivité de l'ARN, Institut de Biologie Moléculaire et Cellulaire du CNRS - UPR 9002, Strasbourg, France
| | - Maria Ericsson
- Electron Microscopy Facility, Harvard Medical School, Boston, USA
| | | | - Ralf Bartenschlager
- Department of Infectious Diseases, Molecular Virology, Heidelberg University, Heidelberg, Germany.,German Centre for Infection Research, Heidelberg University, Heidelberg, Germany
| | - Patrick Pessaux
- Institut National de la Santé et de la Recherche Médicale, U1110, Strasbourg, France.,Université de Strasbourg, Strasbourg, France.,Pôle Hépato-Digestif, Institut Hopitalo-Universitaire, Hôpitaux Universitaires de Strasbourg, Strasbourg, France
| | | | | | - Pascal Villa
- Université de Strasbourg, Strasbourg, France.,Plateforme de Chimie Biologique Intégrative de Strasbourg, UMS 3286 CNRS-UdS & FMTS, Illkirch, France
| | - Lars Kaderali
- Institute for Medical Informatics and Biometry, Medical Faculty, Technische Universität Dresden, Dresden, Germany
| | - Sébastien Pfeffer
- Université de Strasbourg, Strasbourg, France.,Architecture et Réactivité de l'ARN, Institut de Biologie Moléculaire et Cellulaire du CNRS - UPR 9002, Strasbourg, France
| | - Markus H Heim
- Department of Biomedicine, Hepatology Laboratory, University of Basel, Basel, Switzerland
| | - Michel Neunlist
- Institut National de la Santé et de la Recherche Médicale, U913, Nantes, France.,Université de Nantes, Nantes, France.,Institut des Maladies de l'Appareil Digestif, CHU Nantes, Hôpital Hôtel-Dieu, Nantes, France
| | - Mirjam B Zeisel
- Institut National de la Santé et de la Recherche Médicale, U1110, Strasbourg, France.,Université de Strasbourg, Strasbourg, France
| | - Maura Dandri
- I. Department of Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Jane A McKeating
- Hepatitis C Research Group, Institute for Biomedical Research, University of Birmingham, Birmingham, United Kingdom
| | - Eric Robinet
- Institut National de la Santé et de la Recherche Médicale, U1110, Strasbourg, France.,Université de Strasbourg, Strasbourg, France
| | - Thomas F Baumert
- Institut National de la Santé et de la Recherche Médicale, U1110, Strasbourg, France.,Université de Strasbourg, Strasbourg, France.,Pôle Hépato-Digestif, Institut Hopitalo-Universitaire, Hôpitaux Universitaires de Strasbourg, Strasbourg, France
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50
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Meyer K, Kwon YC, Liu S, Hagedorn CH, Ray RB, Ray R. Interferon-α inducible protein 6 impairs EGFR activation by CD81 and inhibits hepatitis C virus infection. Sci Rep 2015; 5:9012. [PMID: 25757571 PMCID: PMC4355636 DOI: 10.1038/srep09012] [Citation(s) in RCA: 51] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/23/2014] [Accepted: 02/16/2015] [Indexed: 02/06/2023] Open
Abstract
Viral entry requires co-operative interactions of several host cell factors. Interferon (IFN) and the IFN-stimulated genes (ISGs) play a central role in antiviral responses against hepatitis C virus (HCV) infection. We examined the effect of interferon-α inducible protein 6 (IFI6) against HCV infection in human hepatoma cells. HCV RNA level or infectious foci were inhibited significantly by ectopic expression of IFI6. IFI6 impaired CD81 co-localization with claudin-1 (CLDN1) upon HCV infection or CD81 cross-linking by specific antibody. Activation of epidermal growth factor receptor (EGFR), a co-factor involved in CD81/CLDN1 interactions, was reduced in IFI6 expressing cells in response to HCV infection or CD81 cross linking by antibody, but not by treatment with EGF. Taken together, the results from our study support a model where IFI6 inhibits HCV entry by impairing EGFR mediated CD81/CLDN1 interactions. This may be relevant to other virus entry processes employing EGFR.
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Affiliation(s)
- Keith Meyer
- Department of Internal Medicine, Saint Louis University
| | | | - Shuanghu Liu
- Department of Medicinal Chemistry, College of Pharmacy, University of Utah
| | - Curt H Hagedorn
- 1] Department of Medicine and Genetics, University of Arkansas for Medical Sciences [2] The Central Arkansas Veterans Healthcare System
| | - Ratna B Ray
- 1] Department of Internal Medicine, Saint Louis University [2] Department of Pathology, Saint Louis University
| | - Ranjit Ray
- 1] Department of Internal Medicine, Saint Louis University [2] Department of Molecular Microbiology &Immunology, Saint Louis University
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