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Misceo D, Mocciaro G, D'Amore S, Vacca M. Diverting hepatic lipid fluxes with lifestyles revision and pharmacological interventions as a strategy to tackle steatotic liver disease (SLD) and hepatocellular carcinoma (HCC). Nutr Metab (Lond) 2024; 21:112. [PMID: 39716321 DOI: 10.1186/s12986-024-00871-3] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/22/2024] [Accepted: 11/13/2024] [Indexed: 12/25/2024] Open
Abstract
Steatotic liver disease (SLD) and Hepatocellular Carcinoma (HCC) are characterised by a substantial rewiring of lipid fluxes caused by systemic metabolic unbalances and/or disrupted intracellular metabolic pathways. SLD is a direct consequence of the interaction between genetic predisposition and a chronic positive energy balance affecting whole-body energy homeostasis and the function of metabolically-competent organs. In this review, we discuss how the impairment of the cross-talk between peripheral organs and the liver stalls glucose and lipid metabolism, leading to unbalances in hepatic lipid fluxes that promote hepatic fat accumulation. We also describe how prolonged metabolic stress builds up toxic lipid species in the liver, and how lipotoxicity and metabolic disturbances drive disease progression by promoting a chronic activation of wound healing, leading to fibrosis and HCC. Last, we provide a critical overview of current state of the art (pre-clinical and clinical evidence) regarding mechanisms of action and therapeutic efficacy of candidate SLD treatment options, and their potential to interfere with SLD/HCC pathophysiology by diverting lipids away from the liver therefore improving metabolic health.
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Affiliation(s)
- Davide Misceo
- Department of Interdisciplinary Medicine, Clinica Medica "C. Frugoni", "Aldo Moro" University of Bari, Piazza Giulio Cesare 11, 70124, Bari, Italy
| | - Gabriele Mocciaro
- Roger Williams Institute of Liver Studies, Foundation for Liver Research, London, SE5 9NT, UK
| | - Simona D'Amore
- Department of Precision and Regenerative Medicine and Ionian Area (DiMePRe-J), Clinica Medica "G. Baccelli", "Aldo Moro" University of Bari, 70124, Bari, Italy.
| | - Michele Vacca
- Department of Interdisciplinary Medicine, Clinica Medica "C. Frugoni", "Aldo Moro" University of Bari, Piazza Giulio Cesare 11, 70124, Bari, Italy.
- Roger Williams Institute of Liver Studies, Foundation for Liver Research, London, SE5 9NT, UK.
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2
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Rani A, Stadler JT, Marsche G. HDL-based therapeutics: A promising frontier in combating viral and bacterial infections. Pharmacol Ther 2024; 260:108684. [PMID: 38964560 DOI: 10.1016/j.pharmthera.2024.108684] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/26/2024] [Revised: 06/03/2024] [Accepted: 07/01/2024] [Indexed: 07/06/2024]
Abstract
Low levels of high-density lipoprotein (HDL) and impaired HDL functionality have been consistently associated with increased susceptibility to infection and its serious consequences. This has been attributed to the critical role of HDL in maintaining cellular lipid homeostasis, which is essential for the proper functioning of immune and structural cells. HDL, a multifunctional particle, exerts pleiotropic effects in host defense against pathogens. It functions as a natural nanoparticle, capable of sequestering and neutralizing potentially harmful substances like bacterial lipopolysaccharides. HDL possesses antiviral activity, preventing viruses from entering or fusing with host cells, thereby halting their replication cycle. Understanding the complex relationship between HDL and the immune system may reveal innovative targets for developing new treatments to combat infectious diseases and improve patient outcomes. This review aims to emphasize the role of HDL in influencing the course of bacterial and viral infections and its and its therapeutic potential.
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Affiliation(s)
- Alankrita Rani
- Division of Pharmacology, Otto Loewi Research Center, Medical University of Graz, Neue Stiftingtalstrasse 6, 8010 Graz, Styria, Austria
| | - Julia T Stadler
- Division of Pharmacology, Otto Loewi Research Center, Medical University of Graz, Neue Stiftingtalstrasse 6, 8010 Graz, Styria, Austria
| | - Gunther Marsche
- Division of Pharmacology, Otto Loewi Research Center, Medical University of Graz, Neue Stiftingtalstrasse 6, 8010 Graz, Styria, Austria; BioTechMed-Graz, Mozartgasse 12/II, 8010 Graz, Styria, Austria.
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3
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Pewkliang Y, Thongsri P, Suthivanich P, Thongbaiphet N, Keatkla J, Pasomsub E, Anurathapan U, Borwornpinyo S, Wongkajornsilp A, Hongeng S, Sa-Ngiamsuntorn K. Immortalized hepatocyte-like cells: A competent hepatocyte model for studying clinical HCV isolate infection. PLoS One 2024; 19:e0303265. [PMID: 38739590 PMCID: PMC11090328 DOI: 10.1371/journal.pone.0303265] [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: 10/31/2023] [Accepted: 04/23/2024] [Indexed: 05/16/2024] Open
Abstract
More than 58 million individuals worldwide are inflicted with chronic HCV. The disease carries a high risk of end stage liver disease, i.e., cirrhosis and hepatocellular carcinoma. Although direct-acting antiviral agents (DAAs) have revolutionized therapy, the emergence of drug-resistant strains has become a growing concern. Conventional cellular models, Huh7 and its derivatives were very permissive to only HCVcc (JFH-1), but not HCV clinical isolates. The lack of suitable host cells had hindered comprehensive research on patient-derived HCV. Here, we established a novel hepatocyte model for HCV culture to host clinically pan-genotype HCV strains. The immortalized hepatocyte-like cell line (imHC) derived from human mesenchymal stem cell carries HCV receptors and essential host factors. The imHC outperformed Huh7 as a host for HCV (JFH-1) and sustained the entire HCV life cycle of pan-genotypic clinical isolates. We analyzed the alteration of host markers (i.e., hepatic markers, cellular innate immune response, and cell apoptosis) in response to HCV infection. The imHC model uncovered the underlying mechanisms governing the action of IFN-α and the activation of sofosbuvir. The insights from HCV-cell culture model hold promise for understanding disease pathogenesis and novel anti-HCV development.
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Affiliation(s)
- Yongyut Pewkliang
- Faculty of Medicine Ramathibodi Hospital, Program in Translational Medicine, Mahidol University, Rama VI Road, Rajathevi, Bangkok, Thailand
| | - Piyanoot Thongsri
- Faculty of Medicine Ramathibodi Hospital, Program in Translational Medicine, Mahidol University, Rama VI Road, Rajathevi, Bangkok, Thailand
| | - Phichaya Suthivanich
- Faculty of Science, Excellent Center for Drug Discovery, Mahidol University, Rama VI Road, Rajathevi, Bangkok, Thailand
| | - Nipa Thongbaiphet
- Faculty of Medicine Ramathibodi Hospital, Department of Pathology, Virology Laboratory, Mahidol University, Rajathevi, Bangkok, Thailand
| | - Jiraporn Keatkla
- Faculty of Medicine Ramathibodi Hospital, Department of Pathology, Virology Laboratory, Mahidol University, Rajathevi, Bangkok, Thailand
| | - Ekawat Pasomsub
- Faculty of Medicine Ramathibodi Hospital, Department of Pathology, Virology Laboratory, Mahidol University, Rajathevi, Bangkok, Thailand
| | - Usanarat Anurathapan
- Faculty of Medicine Ramathibodi Hospital, Department of Pediatrics, Mahidol University, Rajathevi, Bangkok, Thailand
| | - Suparerk Borwornpinyo
- Faculty of Science, Excellent Center for Drug Discovery, Mahidol University, Rama VI Road, Rajathevi, Bangkok, Thailand
- Faculty of Science, Department of Biotechnology, Mahidol University, Rajathevi, Bangkok, Thailand
| | - Adisak Wongkajornsilp
- Faculty of Medicine Siriraj Hospital, Department of Pharmacology, Mahidol University, Bangkok, Thailand
| | - Suradej Hongeng
- Faculty of Medicine Ramathibodi Hospital, Department of Pediatrics, Mahidol University, Rajathevi, Bangkok, Thailand
| | - Khanit Sa-Ngiamsuntorn
- Faculty of Pharmacy, Department of Biochemistry, Mahidol University, Rajathevi, Bangkok, Thailand
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Matthaei A, Joecks S, Frauenstein A, Bruening J, Bankwitz D, Friesland M, Gerold G, Vieyres G, Kaderali L, Meissner F, Pietschmann T. Landscape of protein-protein interactions during hepatitis C virus assembly and release. Microbiol Spectr 2024; 12:e0256222. [PMID: 38230952 PMCID: PMC10846047 DOI: 10.1128/spectrum.02562-22] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/05/2022] [Accepted: 10/11/2023] [Indexed: 01/18/2024] Open
Abstract
Assembly of infectious hepatitis C virus (HCV) particles requires multiple cellular proteins including for instance apolipoprotein E (ApoE). To describe these protein-protein interactions, we performed an affinity purification mass spectrometry screen of HCV-infected cells. We used functional viral constructs with epitope-tagged envelope protein 2 (E2), protein (p) 7, or nonstructural protein 4B (NS4B) as well as cells expressing a tagged variant of ApoE. We also evaluated assembly stage-dependent remodeling of protein complexes by using viral mutants carrying point mutations abrogating particle production at distinct steps of the HCV particle production cascade. Five ApoE binding proteins, 12 p7 binders, 7 primary E2 interactors, and 24 proteins interacting with NS4B were detected. Cell-derived PREB, STT3B, and SPCS2 as well as viral NS2 interacted with both p7 and E2. Only GTF3C3 interacted with E2 and NS4B, highlighting that HCV assembly and replication complexes exhibit largely distinct interactomes. An HCV core protein mutation, preventing core protein decoration of lipid droplets, profoundly altered the E2 interactome. In cells replicating this mutant, E2 interactions with HSPA5, STT3A/B, RAD23A/B, and ZNF860 were significantly enhanced, suggesting that E2 protein interactions partly depend on core protein functions. Bioinformatic and functional studies including STRING network analyses, RNA interference, and ectopic expression support a role of Rad23A and Rad23B in facilitating HCV infectious virus production. Both Rad23A and Rad23B are involved in the endoplasmic reticulum (ER)-associated protein degradation (ERAD). Collectively, our results provide a map of host proteins interacting with HCV assembly proteins, and they give evidence for the involvement of ER protein folding machineries and the ERAD pathway in the late stages of the HCV replication cycle.IMPORTANCEHepatitis C virus (HCV) establishes chronic infections in the majority of exposed individuals. This capacity likely depends on viral immune evasion strategies. One feature likely contributing to persistence is the formation of so-called lipo-viro particles. These peculiar virions consist of viral structural proteins and cellular lipids and lipoproteins, the latter of which aid in viral attachment and cell entry and likely antibody escape. To learn about how lipo-viro particles are coined, here, we provide a comprehensive overview of protein-protein interactions in virus-producing cells. We identify numerous novel and specific HCV E2, p7, and cellular apolipoprotein E-interacting proteins. Pathway analyses of these interactors show that proteins participating in processes such as endoplasmic reticulum (ER) protein folding, ER-associated protein degradation, and glycosylation are heavily engaged in virus production. Moreover, we find that the proteome of HCV replication sites is distinct from the assembly proteome, suggesting that transport process likely shuttles viral RNA to assembly sites.
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Affiliation(s)
- Alina Matthaei
- Institute of Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, Hannover, Lower Saxony, Germany
| | - Sebastian Joecks
- Institute of Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, Hannover, Lower Saxony, Germany
| | - Annika Frauenstein
- RG Experimental Systems Immunology, Max-Planck Institute for Biochemistry, Planegg, Bavaria, Germany
| | - Janina Bruening
- Institute of Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, Hannover, Lower Saxony, Germany
| | - Dorothea Bankwitz
- Institute of Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, Hannover, Lower Saxony, Germany
| | - Martina Friesland
- Institute of Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, Hannover, Lower Saxony, Germany
| | - Gisa Gerold
- Institute of Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, Hannover, Lower Saxony, Germany
- Department of Biochemistry & Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hannover, Hannover, Lower Saxony, Germany
- Department of Clinical Microbiology, Virology, Umeå University, Umeå, Sweden
- Wallenberg Centre for Molecular Medicine (WCMM), Umeå University, Umeå, Sweden
| | - Gabrielle Vieyres
- Institute of Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, Hannover, Lower Saxony, Germany
- Junior Research Group “Cell Biology of RNA Viruses,” Leibniz Institute of Experimental Virology, Hamburg, Germany
| | - Lars Kaderali
- Institute of Bioinformatics, University Medicine Greifswald, Greifswald, Germany
| | - Felix Meissner
- RG Experimental Systems Immunology, Max-Planck Institute for Biochemistry, Planegg, Bavaria, Germany
- Systems Immunology and Proteomics, Institute of Innate Immunity, Medical Faculty, University of Bonn, Bonn, Germany
| | - Thomas Pietschmann
- Institute of Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, Hannover, Lower Saxony, Germany
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Lee WP, Liao SX, Huang YH, Hou MC, Lan KH. Akt1 is involved in HCV release by promoting endoplasmic reticulum-to-endosome transition of infectious virions. Life Sci 2024; 338:122412. [PMID: 38191051 DOI: 10.1016/j.lfs.2024.122412] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2023] [Revised: 12/25/2023] [Accepted: 01/02/2024] [Indexed: 01/10/2024]
Abstract
AIMS Hepatitis C virus (HCV) relies on the viral and host factors to complete its life cycle. It has evolved to profit from Akt activation at some stage in its life cycle through various mechanisms, notably by activating lipogenesis, which is crucial for infectious virions production. MATERIALS AND METHODS By employing an Akt-specific inhibitor, the impact of Akt on intracellular and extracellular infectivity was investigated. To ascertain the role of Akt in the HCV life cycle, the two-part cell culture-derived HCV infection protocol utilizing Akt1 small interfering RNAs (siRNAs) was implemented. The impact of Akt1 on intracellular HCV transition was determined using membrane flotation assay and proximity ligation assay coupled with Anti-Rab7 immunoprecipitation and immunofluorescence. KEY FINDINGS Akt1 silencing reduced infectious virions release to a degree comparable to that of ApoE, a host component involved in the HCV assembly and release, suggesting Akt1 was critical in the late stage of the HCV life cycle. Extracellular infectivity of HCV was inhibited by brefeldin A, and the inhibitory effect was augmented by Akt1 silencing and partially restored by ectopic Akt1 expression. Immunofluorescence revealed that Akt1 inhibition suppressed the interaction between HCV core protein and lipid droplet. Akt1 silencing impeded the transition of HCV from the endoplasmic reticulum to the endosome and hence inhibited the secretion of HCV infectious virions from the late endosome. SIGNIFICANCE Our study demonstrates that Akt1 has an impact on the lipogenesis pathway and plays a critical role in the assembly and secretion of infectious HCV.
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Affiliation(s)
- Wei-Ping Lee
- Department of Medical Research, Taipei Veterans General Hospital, Taipei, Taiwan; Institute of Biochemistry and Molecular Biology, School of Life Sciences, National Yang Ming Chiao Tung University, Taipei, Taiwan
| | - Shi-Xian Liao
- Division of Gastroenterology and Hepatology, Department of Medicine, Taipei Veterans General Hospital, Taipei, Taiwan
| | - Yi-Hsiang Huang
- Division of Gastroenterology and Hepatology, Department of Medicine, Taipei Veterans General Hospital, Taipei, Taiwan; School of Medicine, College of Medicine, National Yang Ming Chiao Tung University, Taipei, Taiwan; Institute of Clinical Medicine, College of Medicine, National Yang Ming Chiao Tung University, Taipei, Taiwan
| | - Ming-Chih Hou
- Division of Gastroenterology and Hepatology, Department of Medicine, Taipei Veterans General Hospital, Taipei, Taiwan; School of Medicine, College of Medicine, National Yang Ming Chiao Tung University, Taipei, Taiwan
| | - Keng-Hsin Lan
- Division of Gastroenterology and Hepatology, Department of Medicine, Taipei Veterans General Hospital, Taipei, Taiwan; School of Medicine, College of Medicine, National Yang Ming Chiao Tung University, Taipei, Taiwan; Institute of Pharmacology, College of Medicine, National Yang Ming Chiao Tung University, Taipei, Taiwan.
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6
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Abstract
The steatotic diseases of metabolic dysfunction-associated steatotic liver disease (MASLD), alcohol-associated liver disease (ALD), and chronic hepatitis C (HCV) account for the majority of liver disease prevalence, morbidity, and mortality worldwide. While these diseases have distinct pathogenic and clinical features, dysregulated lipid droplet (LD) organelle biology represents a convergence of pathogenesis in all three. With increasing understanding of hepatocyte LD biology, we now understand the roles of LD proteins involved in these diseases but also how genetics modulate LD biology to either exacerbate or protect against the phenotypes associated with steatotic liver diseases. Here, we review the history of the LD organelle and its biogenesis and catabolism. We also review how this organelle is critical not only for the steatotic phenotype of liver diseases but also for their advanced phenotypes. Finally, we summarize the latest attempts and challenges of leveraging LD biology for therapeutic gain in steatotic diseases. In conclusion, the study of dysregulated LD biology may lead to novel therapeutics for the prevention of disease progression in the highly prevalent steatotic liver diseases of MASLD, ALD, and HCV.
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Affiliation(s)
- Joseph L Dempsey
- Division of Gastroenterology, Department of Medicine, School of Medicine, University of Washington, Seattle, Washington
| | - George N Ioannou
- Division of Gastroenterology, Department of Medicine, School of Medicine, University of Washington, Seattle, Washington
- Division of Gastroenterology, Veterans Affairs Puget Sound Healthcare System Seattle, Washington
| | - Rotonya M Carr
- Division of Gastroenterology, Department of Medicine, School of Medicine, University of Washington, Seattle, Washington
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Granier C, Toesca J, Mialon C, Ritter M, Freitas N, Boson B, Pécheur EI, Cosset FL, Denolly S. Low-density hepatitis C virus infectious particles are protected from oxidation by secreted cellular proteins. mBio 2023; 14:e0154923. [PMID: 37671888 PMCID: PMC10653866 DOI: 10.1128/mbio.01549-23] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/17/2023] [Accepted: 07/04/2023] [Indexed: 09/07/2023] Open
Abstract
IMPORTANCE Assessments of viral stability on surfaces or in body fluids under different environmental conditions and/or temperatures are often performed, as they are key to understanding the routes and parameters of viral transmission and to providing clues on the epidemiology of infections. However, for most viruses, the mechanisms of inactivation vs stability of viral particles remain poorly defined. Although they are structurally diverse, with different compositions, sizes, and shapes, enveloped viruses are generally less stable than non-enveloped viruses, pointing out the role of envelopes themselves in virus lability. In this report, we investigated the properties of hepatitis C virus (HCV) particles with regards to their stability. We found that, compared to alternative enveloped viruses such as Dengue virus (DENV), severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), hepatitis delta virus (HDV), and Crimean-Congo hemorrhagic fever virus (CCHFV) that infect the liver, HCV particles are intrinsically labile. We determined the mechanisms that drastically alter their specific infectivity through oxidation of their lipids, and we highlighted that they are protected from lipid oxidation by secreted cellular proteins, which can protect their membrane fusion capacity and overall infectivity.
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Affiliation(s)
- Christelle Granier
- CIRI – Centre International de Recherche en Infectiologie, Univ. Lyon, Université Claude Bernard Lyon 1, Inserm, U1111, CNRS, UMR5308 ENS de Lyon, Lyon, France
| | - Johan Toesca
- CIRI – Centre International de Recherche en Infectiologie, Univ. Lyon, Université Claude Bernard Lyon 1, Inserm, U1111, CNRS, UMR5308 ENS de Lyon, Lyon, France
| | - Chloé Mialon
- CIRI – Centre International de Recherche en Infectiologie, Univ. Lyon, Université Claude Bernard Lyon 1, Inserm, U1111, CNRS, UMR5308 ENS de Lyon, Lyon, France
| | - Maureen Ritter
- CIRI – Centre International de Recherche en Infectiologie, Univ. Lyon, Université Claude Bernard Lyon 1, Inserm, U1111, CNRS, UMR5308 ENS de Lyon, Lyon, France
| | - Natalia Freitas
- CIRI – Centre International de Recherche en Infectiologie, Univ. Lyon, Université Claude Bernard Lyon 1, Inserm, U1111, CNRS, UMR5308 ENS de Lyon, Lyon, France
| | - Bertrand Boson
- CIRI – Centre International de Recherche en Infectiologie, Univ. Lyon, Université Claude Bernard Lyon 1, Inserm, U1111, CNRS, UMR5308 ENS de Lyon, Lyon, France
| | - Eve-Isabelle Pécheur
- Centre Léon Bérard, Centre de Recherche en Cancérologie de Lyon, CNRS 5286, Inserm U1052, Université Claude Bernard Lyon 1, Lyon, France
| | - François-Loïc Cosset
- CIRI – Centre International de Recherche en Infectiologie, Univ. Lyon, Université Claude Bernard Lyon 1, Inserm, U1111, CNRS, UMR5308 ENS de Lyon, Lyon, France
| | - Solène Denolly
- CIRI – Centre International de Recherche en Infectiologie, Univ. Lyon, Université Claude Bernard Lyon 1, Inserm, U1111, CNRS, UMR5308 ENS de Lyon, Lyon, France
- Department of Infectious Diseases, Molecular Virology, Heidelberg University, Heidelberg, Germany
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8
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Avula K, Singh B, Samantaray S, Syed GH. The Early Secretory Pathway Is Crucial for Multiple Aspects of the Hepatitis C Virus Life Cycle. J Virol 2023:e0018023. [PMID: 37338368 PMCID: PMC10373535 DOI: 10.1128/jvi.00180-23] [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: 06/01/2023] [Accepted: 06/02/2023] [Indexed: 06/21/2023] Open
Abstract
Although most of the early events of the hepatitis C virus (HCV) life cycle are well characterized, our understanding of HCV egress is still unclear. Some reports implicate the conventional endoplasmic reticulum (ER)-Golgi route, while some propose noncanonical secretory routes. Initially, the envelopment of HCV nucleocapsid occurs by budding into the ER lumen. Subsequently, the HCV particle exit from the ER is assumed to be mediated by coat protein complex II (COPII) vesicles. COPII vesicle biogenesis also involves the recruitment of cargo to the site of vesicle biogenesis via interaction with COPII inner coat proteins. We investigated the modulation and the specific role of the individual components of the early secretory pathway in HCV egress. We observed that HCV inhibits cellular protein secretion and triggers the reorganization of the ER exit sites and ER-Golgi intermediate compartments (ERGIC). Gene-specific knockdown of the components of this pathway such as SEC16A, TFG, ERGIC-53, and COPII coat proteins demonstrated the functional significance of these components and the distinct role played by these proteins in various aspects of the HCV life cycle. SEC16A is essential for multiple steps in the HCV life cycle, whereas TFG is specifically involved in HCV egress and ERGIC-53 is crucial for HCV entry. Overall, our study establishes that the components of the early secretory pathway are essential for HCV propagation and emphasize the importance of the ER-Golgi secretory route in this process. Surprisingly, these components are also required for the early stages of the HCV life cycle due to their role in overall intracellular trafficking and homeostasis of the cellular endomembrane system. IMPORTANCE The virus life cycle involves entry into the host, replication of the genome, assembly of infectious progeny, and their subsequent release. Different aspects of the HCV life cycle, including entry, genome replication, and assembly, are well characterized; however, our understanding of the HCV release is still not clear and subject to debate due to varied findings. Here, we attempted to address this controversy and enhance our understanding of HCV egress by evaluating the role of the different components of the early secretory pathway in the HCV life cycle. To our surprise, we found that the components of the early secretory pathway are not only essential for HCV release but also contribute to many other earlier events of the HCV life cycle. This study emphasizes the importance of the early secretory pathway for the establishment of productive HCV infection in hepatocytes.
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Affiliation(s)
- Kiran Avula
- Institute of Life Sciences, Bhubaneswar, Odisha, India
- Regional Centre for Biotechnology, Faridabad, Delhi, India
| | - Bharati Singh
- Institute of Life Sciences, Bhubaneswar, Odisha, India
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9
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Gong Z, Yan Z, Liu W, Luo B. Oncogenic viruses and host lipid metabolism: a new perspective. J Gen Virol 2023; 104. [PMID: 37279154 DOI: 10.1099/jgv.0.001861] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 06/08/2023] Open
Abstract
As noncellular organisms, viruses do not have their own metabolism and rely on the metabolism of host cells to provide energy and metabolic substances for their life cycles. Increasing evidence suggests that host cells infected with oncogenic viruses have dramatically altered metabolic requirements and that oncogenic viruses produce substances used for viral replication and virion production by altering host cell metabolism. We focused on the processes by which oncogenic viruses manipulate host lipid metabolism and the lipid metabolism disorders that occur in oncogenic virus-associated diseases. A deeper understanding of viral infections that cause changes in host lipid metabolism could help with the development of new antiviral agents as well as potential new therapeutic targets.
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Affiliation(s)
- Zhiyuan Gong
- Department of Pathogenic Biology, School of Basic Medicine, Qingdao University, Qingdao 266071, PR China
| | - Zhiyong Yan
- Department of Pathogenic Biology, School of Basic Medicine, Qingdao University, Qingdao 266071, PR China
| | - Wen Liu
- Department of Pathogenic Biology, School of Basic Medicine, Qingdao University, Qingdao 266071, PR China
| | - Bing Luo
- Department of Pathogenic Biology, School of Basic Medicine, Qingdao University, Qingdao 266071, PR China
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10
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Abomughaid M, Tay ESE, Pickford R, Malladi C, Read SA, Coorssen JR, Gloss BS, George J, Douglas MW. PEMT Mediates Hepatitis C Virus-Induced Steatosis, Explains Genotype-Specific Phenotypes and Supports Virus Replication. Int J Mol Sci 2023; 24:ijms24108781. [PMID: 37240132 DOI: 10.3390/ijms24108781] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/14/2023] [Revised: 05/05/2023] [Accepted: 05/11/2023] [Indexed: 05/28/2023] Open
Abstract
The hepatitis C virus (HCV) relies on cellular lipid pathways for virus replication and also induces liver steatosis, but the mechanisms involved are not clear. We performed a quantitative lipidomics analysis of virus-infected cells by combining high-performance thin-layer chromatography (HPTLC) and mass spectrometry, using an established HCV cell culture model and subcellular fractionation. Neutral lipid and phospholipids were increased in the HCV-infected cells; in the endoplasmic reticulum there was an ~four-fold increase in free cholesterol and an ~three-fold increase in phosphatidyl choline (p < 0.05). The increase in phosphatidyl choline was due to the induction of a non-canonical synthesis pathway involving phosphatidyl ethanolamine transferase (PEMT). An HCV infection induced expression of PEMT while knocking down PEMT with siRNA inhibited virus replication. As well as supporting virus replication, PEMT mediates steatosis. Consistently, HCV induced the expression of the pro-lipogenic genes SREBP 1c and DGAT1 while inhibiting the expression of MTP, promoting lipid accumulation. Knocking down PEMT reversed these changes and reduced the lipid content in virus-infected cells. Interestingly, PEMT expression was over 50% higher in liver biopsies from people infected with the HCV genotype 3 than 1, and three times higher than in people with chronic hepatitis B, suggesting that this may account for genotype-dependent differences in the prevalence of hepatic steatosis. PEMT is a key enzyme for promoting the accumulation of lipids in HCV-infected cells and supports virus replication. The induction of PEMT may account for virus genotype specific differences in hepatic steatosis.
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Affiliation(s)
- Mosleh Abomughaid
- Storr Liver Centre, The Westmead Institute for Medical Research, The University of Sydney and Westmead Hospital, Sydney, NSW 2145, Australia
| | - Enoch S E Tay
- Storr Liver Centre, The Westmead Institute for Medical Research, The University of Sydney and Westmead Hospital, Sydney, NSW 2145, Australia
| | - Russell Pickford
- Bioanalytical Mass Spectrometry Facility, Mark Wainright Analytical Centre, University of New South Wales, Sydney, NSW 2052, Australia
| | - Chandra Malladi
- Department of Molecular Physiology, School of Medicine, Western Sydney University, Sydney, NSW 2751, Australia
| | - Scott A Read
- Storr Liver Centre, The Westmead Institute for Medical Research, The University of Sydney and Westmead Hospital, Sydney, NSW 2145, Australia
- Blacktown Clinical School, Western Sydney University and Blacktown Hospital, Sydney, NSW 2751, Australia
| | - Jens R Coorssen
- Department of Molecular Physiology, School of Medicine, Western Sydney University, Sydney, NSW 2751, Australia
- Department of Biological Sciences, Faculty of Mathematics and Science, Brock University, St. Catharines, ON L2S 3A1, Canada
| | - Brian S Gloss
- Westmead Research Hub, Westmead Institute for Medical Research, Sydney, NSW 2145, Australia
| | - Jacob George
- Storr Liver Centre, The Westmead Institute for Medical Research, The University of Sydney and Westmead Hospital, Sydney, NSW 2145, Australia
| | - Mark W Douglas
- Storr Liver Centre, The Westmead Institute for Medical Research, The University of Sydney and Westmead Hospital, Sydney, NSW 2145, Australia
- Centre for Infectious Diseases and Microbiology, Sydney Infectious Diseases Institute, The University of Sydney at Westmead Hospital, Sydney, NSW 2145, Australia
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11
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Vieyres G, Pietschmann T. The role of human lipoproteins for hepatitis C virus persistence. Curr Opin Virol 2023; 60:101327. [PMID: 37031484 DOI: 10.1016/j.coviro.2023.101327] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/02/2022] [Revised: 01/23/2023] [Accepted: 03/05/2023] [Indexed: 04/11/2023]
Abstract
Hepatitis C virus (HCV) is a hepatotropic virus that establishes a chronic infection in most individuals. Effective treatments are available; however, many patients are not aware of their infection. Consequently, they do not receive treatment and HCV transmission remains high, particularly among groups at high risk of exposure such as people who inject intravenous drugs. A prophylactic vaccine may reduce HCV transmission, but is currently not available. HCV has evolved immune evasion strategies, which facilitate persistence and complicate development of a protective vaccine. The peculiar association of HCV particles with human lipoproteins is thought to facilitate evasion from humoral immune response and viral homing to liver cells. A better understanding of these aspects provides the basis for development of protective vaccination strategies. Here, we review key information about the composition of HCV particles, the mechanisms mediating lipoprotein incorporation, and the functional consequences of this interaction.
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Affiliation(s)
- Gabrielle Vieyres
- Leibniz Institute of Virology, Hamburg, Germany; Integrative Analysis of Pathogen-Induced Compartments, Leibniz ScienceCampus InterACt, Hamburg, 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, Hanover, Germany.
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12
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Awadh AA. The Role of Cytosolic Lipid Droplets in Hepatitis C Virus Replication, Assembly, and Release. BIOMED RESEARCH INTERNATIONAL 2023; 2023:5156601. [PMID: 37090186 PMCID: PMC10121354 DOI: 10.1155/2023/5156601] [Citation(s) in RCA: 12] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 08/20/2022] [Revised: 03/02/2023] [Accepted: 03/09/2023] [Indexed: 04/25/2023]
Abstract
The hepatitis C virus (HCV) causes chronic hepatitis by establishing a persistent infection. Patients with chronic hepatitis frequently develop hepatic cirrhosis, which can lead to liver cancer-the progressive liver damage results from the host's immune response to the unresolved infection. The HCV replication process, including the entry, replication, assembly, and release stages, while the virus circulates in the bloodstream, it is intricately linked to the host's lipid metabolism, including the dynamic of the cytosolic lipid droplets (cLDs). This review article depicts how this interaction regulates viral cell tropism and aids immune evasion by coining viral particle characteristics. cLDs are intracellular organelles that store most of the cytoplasmic components of neutral lipids and are assumed to play an increasingly important role in the pathophysiology of lipid metabolism and host-virus interactions. cLDs are involved in the replication of several clinically significant viruses, where viruses alter the lipidomic profiles of host cells to improve viral life cycles. cLDs are involved in almost every phase of the HCV life cycle. Indeed, pharmacological modulators of cholesterol synthesis and intracellular trafficking, lipoprotein maturation, and lipid signaling molecules inhibit the assembly of HCV virions. Likewise, small-molecule inhibitors of cLD-regulating proteins inhibit HCV replication. Thus, addressing the molecular architecture of HCV replication will aid in elucidating its pathogenesis and devising preventive interventions that impede persistent infection and prevent disease progression. This is possible via repurposing the available therapeutic agents that alter cLDs metabolism. This review highlights the role of cLD in HCV replication.
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Affiliation(s)
- Abdullah A. Awadh
- Department of Basic Medical Sciences, College of Medicine, King Saud bin Abdulaziz University for Health Sciences, Jeddah 21423, Saudi Arabia
- King Abdullah International Medical Research Center, Jeddah 21423, Saudi Arabia
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13
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An Update on the Metabolic Landscape of Oncogenic Viruses. Cancers (Basel) 2022; 14:cancers14235742. [PMID: 36497226 PMCID: PMC9738352 DOI: 10.3390/cancers14235742] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/10/2022] [Revised: 11/10/2022] [Accepted: 11/17/2022] [Indexed: 11/24/2022] Open
Abstract
Viruses play an important role in cancer development as about 12% of cancer types are linked to viral infections. Viruses that induce cellular transformation are known as oncoviruses. Although the mechanisms of viral oncogenesis differ between viruses, all oncogenic viruses share the ability to establish persistent chronic infections with no obvious symptoms for years. During these prolonged infections, oncogenic viruses manipulate cell signaling pathways that control cell cycle progression, apoptosis, inflammation, and metabolism. Importantly, it seems that most oncoviruses depend on these changes for their persistence and amplification. Metabolic changes induced by oncoviruses share many common features with cancer metabolism. Indeed, viruses, like proliferating cancer cells, require increased biosynthetic precursors for virion production, need to balance cellular redox homeostasis, and need to ensure host cell survival in a given tissue microenvironment. Thus, like for cancer cells, viral replication and persistence of infected cells frequently depend on metabolic changes. Here, we draw parallels between metabolic changes observed in cancers or induced by oncoviruses, with a focus on pathways involved in the regulation of glucose, lipid, and amino acids. We describe whether and how oncoviruses depend on metabolic changes, with the perspective of targeting them for antiviral and onco-therapeutic approaches in the context of viral infections.
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14
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Leslie J, Geh D, Elsharkawy AM, Mann DA, Vacca M. Metabolic dysfunction and cancer in HCV: Shared pathways and mutual interactions. J Hepatol 2022; 77:219-236. [PMID: 35157957 DOI: 10.1016/j.jhep.2022.01.029] [Citation(s) in RCA: 22] [Impact Index Per Article: 7.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/31/2021] [Revised: 01/12/2022] [Accepted: 01/31/2022] [Indexed: 12/16/2022]
Abstract
HCV hijacks many host metabolic processes in an effort to aid viral replication. The resulting hepatic metabolic dysfunction underpins many of the hepatic and extrahepatic manifestations of chronic hepatitis C (CHC). However, the natural history of CHC is also substantially influenced by the host metabolic status: obesity, insulin resistance and hepatic steatosis are major determinants of CHC progression toward hepatocellular carcinoma (HCC). Direct-acting antivirals (DAAs) have transformed the treatment and natural history of CHC. While DAA therapy effectively eradicates the virus, the long-lasting overlapping metabolic disease can persist, especially in the presence of obesity, increasing the risk of liver disease progression. This review covers the mechanisms by which HCV tunes hepatic and systemic metabolism, highlighting how systemic metabolic disturbance, lipotoxicity and chronic inflammation favour disease progression and a precancerous niche. We also highlight the therapeutic implications of sustained metabolic dysfunction following sustained virologic response as well as considerations for patients who develop HCC on the background of metabolic dysfunction.
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Affiliation(s)
- Jack Leslie
- Newcastle Fibrosis Research Group, Biosciences Institute, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, UK
| | - Daniel Geh
- Newcastle Fibrosis Research Group, Biosciences Institute, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, UK
| | - Ahmed M Elsharkawy
- Liver Unit, University Hospitals Birmingham NHS Foundation Trust, Queen Elizabeth Hospital, Queen Elizabeth Medical Centre, Birmingham, B15 2TH UK; National Institute for Health Research, Birmingham Biomedical Research Centre at University Hospitals Birmingham NHS Foundation Trust, Birmingham, UK
| | - Derek A Mann
- Newcastle Fibrosis Research Group, Biosciences Institute, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, UK; Department of Gastroenterology and Hepatology, School of Medicine, Koç University, Istanbul, Turkey.
| | - Michele Vacca
- Interdisciplinary Department of Medicine, Università degli Studi di Bari "Aldo Moro", Bari, Italy.
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15
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Sofosbuvir-based direct-acting antivirals and changes in cholesterol and low density lipoprotein-cholesterol. Sci Rep 2022; 12:9942. [PMID: 35705594 PMCID: PMC9200852 DOI: 10.1038/s41598-022-13657-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/21/2021] [Accepted: 05/05/2022] [Indexed: 11/19/2022] Open
Abstract
Worsened lipid profiles were observed in chronic hepatitis C (CHC) patients during direct-acting antivirals (DAAs) treatment, among which combination drugs confounded the effect of individual ingredient on lipid. Tenofovir alafenamide (TAF) also worsened lipid profiles in HIV patients. Structural similarity between sofosbuvir (SOF) and TAF prompted us to investigate rapid increase in total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) in CHC patients treated with SOF-based DAAs. A retrospective study was performed to analyze 487 CHC patients receiving DAAs with SVR12. Relative risks on elevating TC and LDL-C were analyzed by logistic regression to determine SOF-based over non-SOF-based regimens. TC or LDL-C levels at baseline, week-4 and SVR12 were compared by Wilcoxon matched-pairs signed rank test. Week 4 or SVR12 to baseline ratios of serum TC or LDL-C between regimens were compared by Mann–Whitney's test. 487 patients were treated with Harvoni (SOF-based, 206 patients), Epclusa (SOF-based, 124 patients), Maviret (non-SOF-based, 122 patients), or Zepatier (non-SOF-based, 35 patients). At week 4 during drug treatment, Harvoni, Epclusa, and Maviret induced statistically significant elevation of TC and LDL-C, but Zepatier did not. SOF-based regimens had 2.72-fold higher relative risk (RR) causing 10% elevation of TC (95% CI 1.84–4.02, p < 0.001) and 2.04-fold higher RR causing 10% elevation of LDL-C (95% CI 1.39–3.01, p < 0.001) than non-SOF-based DAAs. SOF-based DAAs were associated with significantly larger amplitude of increases in TC and LDL-C than non-SOF-based DAAs during the initial 4 weeks of treatment, but the increases were not sustained to SVR12.
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16
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Poly(rC)-Binding Protein 1 Limits Hepatitis C Virus Virion Assembly and Secretion. Viruses 2022; 14:v14020291. [PMID: 35215884 PMCID: PMC8877974 DOI: 10.3390/v14020291] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/10/2022] [Revised: 01/24/2022] [Accepted: 01/26/2022] [Indexed: 12/16/2022] Open
Abstract
The hepatitis C virus (HCV) co-opts numerous cellular elements, including proteins, lipids, and microRNAs, to complete its viral life cycle. The cellular RNA-binding protein, poly(rC)-binding protein 1 (PCBP1), was previously reported to bind to the 5′ untranslated region (UTR) of the HCV genome; however, its importance in the viral life cycle has remained unclear. Herein, we sought to clarify the role of PCBP1 in the HCV life cycle. Using the HCV cell culture (HCVcc) system, we found that knockdown of endogenous PCBP1 resulted in an overall decrease in viral RNA accumulation, yet resulted in an increase in extracellular viral titers. To dissect PCBP1’s specific role in the HCV life cycle, we carried out assays for viral entry, translation, genome stability, RNA replication, as well as virion assembly and secretion. We found that PCBP1 knockdown did not directly affect viral entry, translation, RNA stability, or RNA replication, but resulted in an overall increase in infectious particle secretion. This increase in virion secretion was evident even when viral RNA synthesis was inhibited, and blocking virus secretion could partially restore the viral RNA accumulation decreased by PCBP1 knockdown. We therefore propose a model where endogenous PCBP1 normally limits virion assembly and secretion, which increases viral RNA accumulation in infected cells by preventing the departure of viral genomes packaged into virions. Overall, our findings improve our understanding of how cellular RNA-binding proteins influence viral genomic RNA utilization during the HCV life cycle.
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17
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Wang YW, Lee WP, Huang YH, Hou MC, Lan KH. Effect of sofosbuvir-based DAAs on changes in lower-density lipoprotein in HCV patients: a systematic review and meta-analysis. BMC Infect Dis 2021; 21:984. [PMID: 34548026 PMCID: PMC8454153 DOI: 10.1186/s12879-021-06657-9] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/16/2021] [Accepted: 09/04/2021] [Indexed: 11/10/2022] Open
Abstract
BACKGROUND Previous studies reported worsened lipid profiles in patients infected with hepatitis C virus (HCV) during direct-acting antivirals (DAAs) treatment. This study aimed to investigate the effect of sofosbuvir (SOF)-based DAAs on changes in low-density lipoprotein (LDL) in HCV patients. METHODS A systematic review of articles published before 31 May 2021 was conducted by searching MEDLINE, Cochrane Library, EMBASE, and CINAHL Plus. Eligible studies were those comparing SOF-based DAAs and non-SOF DAAs for HCV patients and providing numerical data for changes in LDL. Risk of Bias in Non-randomized Studies- of Interventions was used for assessing risk of bias, and meta-analysis was performed for changes in LDL. RESULTS Six studies comprising 1248 patients were included, 848 patients treated with SOF-based DAAs and 400 patients with non-SOF DAAs vs. SOF-based DAAs group had significantly greater increases in LDL from baseline to week 4 than non-SOF DAAs group (P = 0.001). However, changes in LDL from baseline to the end of treatment (P = 0.060), to post-treatment week 12 (P = 0.263), and to post-treatment week 24 (P = 0.319) did not significantly differ between the two groups. Further comparison of SOF/ledipasvir with asunaprevir/daclatasvir revealed a similar trend in changes in LDL. CONCLUSIONS For HCV patients, SOF-based DAA regimens were associated with rapid and significant increases in LDL during the initial 4 weeks of treatment, and the changes did not sustain after the end of treatment. Potential mechanism might be related to the phosphoramidate side chain of SOF.
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Affiliation(s)
- Ying-Wen Wang
- Healthcare and Management Center, Taipei Veterans General Hospital, Taipei, Taiwan, ROC
- School of Medicine, College of Medicine, National Yang Ming Chiao Tung University, Taipei, Taiwan, ROC
| | - Wei-Ping Lee
- Department of Medical Research, Taipei Veterans General Hospital, Taipei, Taiwan, ROC
- Institute of Biochemistry and Molecular Biology, College of Life Sciences, National Yang Ming Chiao Tung University, Taipei, Taiwan, ROC
| | - Yi-Hsiang Huang
- Division of Gastroenterology and Hepatology, Department of Medicine, Taipei Veterans General Hospital, 201, Section 2, Shi-Pai Road, Taipei, 112 Taiwan, ROC
- School of Medicine, College of Medicine, National Yang Ming Chiao Tung University, Taipei, Taiwan, ROC
- Institute of Clinical Medicine, College of Medicine, National Yang Ming Chiao Tung University, Taipei, Taiwan, ROC
| | - Ming-Chih Hou
- Division of Gastroenterology and Hepatology, Department of Medicine, Taipei Veterans General Hospital, 201, Section 2, Shi-Pai Road, Taipei, 112 Taiwan, ROC
- School of Medicine, College of Medicine, National Yang Ming Chiao Tung University, Taipei, Taiwan, ROC
| | - Keng-Hsin Lan
- Division of Gastroenterology and Hepatology, Department of Medicine, Taipei Veterans General Hospital, 201, Section 2, Shi-Pai Road, Taipei, 112 Taiwan, ROC
- School of Medicine, College of Medicine, National Yang Ming Chiao Tung University, Taipei, Taiwan, ROC
- Institute of Pharmacology, College of Medicine, National Yang Ming Chiao Tung University, Taipei, Taiwan, ROC
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18
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Cochard J, Bull-Maurer A, Tauber C, Burlaud-Gaillard J, Mazurier F, Meunier JC, Roingeard P, Chouteau P. Differentiated Cells in Prolonged Hypoxia Produce Highly Infectious Native-Like Hepatitis C Virus Particles. Hepatology 2021; 74:627-640. [PMID: 33665810 DOI: 10.1002/hep.31788] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/08/2020] [Revised: 01/28/2021] [Accepted: 02/05/2021] [Indexed: 01/20/2023]
Abstract
BACKGROUND AND AIMS Standard hepatitis C virus (HCV) cell-culture models present an altered lipid metabolism and thus produce lipid-poor lipoviral particles (LVPs). These models are thereby weakly adapted to explore the complete natural viral life cycle. APPROACH AND RESULTS To overcome these limitations, we used an HCV cell-culture model based on both cellular differentiation and sustained hypoxia to better mimic the host-cell environment. The long-term exposure of Huh7.5 cells to DMSO and hypoxia (1% O2 ) significantly enhanced the expression of major differentiation markers and the cellular hypoxia adaptive response by contrast with undifferentiated and normoxic (21% O2 ) standard conditions. Because hepatocyte-like differentiation and hypoxia are key regulators of intracellular lipid metabolism, we characterized the distribution of lipid droplets (LDs) and demonstrated that experimental cells significantly accumulate larger and more numerous LDs relative to standard cell-culture conditions. An immunocapture (IC) and transmission electron microscopy (TEM) method showed that differentiated and hypoxic Huh7.5 cells produced lipoproteins significantly larger than those produced by standard Huh7.5 cell cultures. The experimental cell culture model is permissive to HCV-Japanese fulminant hepatitis (JFH1) infection and produces very-low-buoyant-density LVPs that are 6-fold more infectious than LVPs formed by standard JFH1-infected Huh7.5 cells. Finally, the IC-TEM approach and antibody-neutralization experiments revealed that LVPs were highly lipidated, had a global ultrastructure and a conformation of the envelope glycoprotein complex E1E2 close to that of the ones circulating in infected individuals. CONCLUSIONS This relevant HCV cell culture model thus mimics the complete native intracellular HCV life cycle and, by extension, can be proposed as a model of choice for studies of other hepatotropic viruses.
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Affiliation(s)
- Jade Cochard
- INSERM U1259Université de Tours and CHRU de ToursToursFrance
| | | | - Clovis Tauber
- UMRS INSERM U1253 Imagerie et cerveauUniversité de ToursToursFrance
| | | | - Frédéric Mazurier
- Université de ToursEquipe Associée 5501CNRS Equipe de Recherche Labellisée 7001LNOx TeamToursFrance
| | | | - Philippe Roingeard
- INSERM U1259Université de Tours and CHRU de ToursToursFrance.,Plate-Forme IBiSA des MicroscopiesUniversité de Tours and CHRU de ToursToursFrance
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19
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Spaziante M, Taliani G, Marchetti G, Tavelli A, Lichtner M, Cingolani A, Cicalini S, Biliotti E, Girardi E, Antinori A, Puoti M, d’Arminio Monforte A, Cozzi-Lepri A. Impact of HCV Eradication on Lipid Metabolism in HIV/HCV Coinfected Patients: Data from ICONA and HepaICONA Foundation Cohort Study. Viruses 2021; 13:v13071402. [PMID: 34372608 PMCID: PMC8310285 DOI: 10.3390/v13071402] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/10/2021] [Revised: 07/13/2021] [Accepted: 07/13/2021] [Indexed: 12/23/2022] Open
Abstract
OBJECTIVES HCV shows complex interactions with lipid metabolism. Our aim was to examine total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) changes in HIV/HCV coinfected patients, after achieving sustained virological response (SVR), according to different HCV genotypes and specific antiretroviral use. METHODS HIV/HCV coinfected patients, enrolled in the ICONA and HepaICONA cohorts, who achieved DAA-driven SVR were included. Paired t-tests were used to examine whether the pre- and post-SVR laboratory value variations were significantly different from zero. ANCOVA regression models were employed to estimate the causal effect of SVR and of PI/r use on lipid changes. The interaction between the effect of eradication and HCV genotype was formally tested. RESULTS six hundred and ninety-nine HIV/HCV coinfected patients were enrolled. After HCV eradication, a significant improvement in liver function occurred, with a significant decrease in AST, ALT, GGT, and total plasmatic bilirubin. TC and LDL-C significantly increased by 21.4 mg/dL and 22.4 mg/dL, respectively (p < 0.001), after SVR, whereas there was no evidence for a change in HDL-C (p = 0.45) and triglycerides (p = 0.49). Notably, the TC and LDL-C increase was higher for participants who were receiving darunavir/ritonavir, and the TC showed a more pronounced increase among HCV genotype 3 patients (interaction-p value = 0.002). CONCLUSIONS complex and rapid changes in TC and LDL-C levels, modulated by HCV genotype and PI/r-based ART combinations, occurred in HIV/HCV coinfected patients after SVR. Further studies are needed to evaluate the clinical impact of these changes on the long-term risk of cardiovascular disease.
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Affiliation(s)
- Martina Spaziante
- Clinical Epidemiology Unit, National Institute for Infectious Diseases Lazzaro Spallanzani IRCCS, 00149 Rome, Italy; (M.S.); (E.G.)
| | - Gloria Taliani
- Department of Translational and Precision Medicine, Sapienza University of Rome, 00185 Rome, Italy;
- Task Force Anti-COVID, AORN San Giuseppe Moscati, 83100 Avellino, Italy
- Correspondence:
| | - Giulia Marchetti
- ASST Santi Paolo e Carlo, Clinic of Infectious and Tropical Diseases, Department of Health Sciences, University of Milan, 20122 Milan, Italy; (G.M.); (A.d.M.)
| | | | - Miriam Lichtner
- Department of Infectious Diseases, La Sapienza University, Polo Pontino, 04100 Latina, Italy;
| | - Antonella Cingolani
- Institute of Clinical Infectious Diseases, Catholic University of the Sacred Heart, Policlinico Agostino Gemelli IRCCS, 00168 Rome, Italy;
| | - Stefania Cicalini
- HIV/AIDS Clinical Unit, National Institute for Infectious Diseases Lazzaro Spallanzani IRCCS, 00149 Rome, Italy; (S.C.); (A.A.)
| | - Elisa Biliotti
- Department of Translational and Precision Medicine, Sapienza University of Rome, 00185 Rome, Italy;
| | - Enrico Girardi
- Clinical Epidemiology Unit, National Institute for Infectious Diseases Lazzaro Spallanzani IRCCS, 00149 Rome, Italy; (M.S.); (E.G.)
| | - Andrea Antinori
- HIV/AIDS Clinical Unit, National Institute for Infectious Diseases Lazzaro Spallanzani IRCCS, 00149 Rome, Italy; (S.C.); (A.A.)
| | - Massimo Puoti
- Division of Infectious Diseases, ASST Grande Ospedale Metropolitano Niguarda, 20162 Milan, Italy;
| | - Antonella d’Arminio Monforte
- ASST Santi Paolo e Carlo, Clinic of Infectious and Tropical Diseases, Department of Health Sciences, University of Milan, 20122 Milan, Italy; (G.M.); (A.d.M.)
| | - Alessandro Cozzi-Lepri
- Centre for Clinical Research, Epidemiology, Modelling and Evaluation (CREME), Institute for Global Health, UCL, London WC1N 1EH, UK;
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20
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LeBlanc EV, Kim Y, Capicciotti CJ, Colpitts CC. Hepatitis C Virus Glycan-Dependent Interactions and the Potential for Novel Preventative Strategies. Pathogens 2021; 10:pathogens10060685. [PMID: 34205894 PMCID: PMC8230238 DOI: 10.3390/pathogens10060685] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/05/2021] [Revised: 05/25/2021] [Accepted: 05/28/2021] [Indexed: 12/15/2022] Open
Abstract
Chronic hepatitis C virus (HCV) infections continue to be a major contributor to liver disease worldwide. HCV treatment has become highly effective, yet there are still no vaccines or prophylactic strategies available to prevent infection and allow effective management of the global HCV burden. Glycan-dependent interactions are crucial to many aspects of the highly complex HCV entry process, and also modulate immune evasion. This review provides an overview of the roles of viral and cellular glycans in HCV infection and highlights glycan-focused advances in the development of entry inhibitors and vaccines to effectively prevent HCV infection.
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Affiliation(s)
- Emmanuelle V. LeBlanc
- Department of Biomedical and Molecular Sciences, Queen’s University, Kingston, ON K7L 3N6, Canada; (E.V.L.); (Y.K.); (C.J.C.)
| | - Youjin Kim
- Department of Biomedical and Molecular Sciences, Queen’s University, Kingston, ON K7L 3N6, Canada; (E.V.L.); (Y.K.); (C.J.C.)
| | - Chantelle J. Capicciotti
- Department of Biomedical and Molecular Sciences, Queen’s University, Kingston, ON K7L 3N6, Canada; (E.V.L.); (Y.K.); (C.J.C.)
- Department of Chemistry, Queen’s University, Kingston, ON K7L 3N6, Canada
- Department of Surgery, Queen’s University, Kingston, ON K7L 3N6, Canada
| | - Che C. Colpitts
- Department of Biomedical and Molecular Sciences, Queen’s University, Kingston, ON K7L 3N6, Canada; (E.V.L.); (Y.K.); (C.J.C.)
- Correspondence:
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21
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Hepatitis C Virus Uses Host Lipids to Its Own Advantage. Metabolites 2021; 11:metabo11050273. [PMID: 33925362 PMCID: PMC8145847 DOI: 10.3390/metabo11050273] [Citation(s) in RCA: 20] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/15/2021] [Revised: 04/11/2021] [Accepted: 04/23/2021] [Indexed: 02/06/2023] Open
Abstract
Lipids and lipoproteins constitute indispensable components for living not only for humans. In the case of hepatitis C virus (HCV), the option of using the products of our lipid metabolism is “to be, or not to be”. On the other hand, HCV infection, which is the main cause of chronic hepatitis, cirrhosis and hepatocellular carcinoma, exerts a profound influence on lipid and lipoprotein metabolism of the host. The consequences of this alternation are frequently observed as hypolipidemia and hepatic steatosis in chronic hepatitis C (CHC) patients. The clinical relevance of these changes reflects the fact that lipids and lipoprotein play a crucial role in all steps of the life cycle of HCV. The virus circulates in the bloodstream as a highly lipidated lipo-viral particle (LVP) that defines HCV hepatotropism. Thus, strict relationships between lipids/lipoproteins and HCV are indispensable for the mechanism of viral entry into hepatocytes, viral replication, viral particles assembly and secretion. The purpose of this review is to summarize the tricks thanks to which HCV utilizes host lipid metabolism to its own advantage.
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22
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Hernandez-Gonzalez M, Larocque G, Way M. Viral use and subversion of membrane organization and trafficking. J Cell Sci 2021; 134:jcs252676. [PMID: 33664154 PMCID: PMC7610647 DOI: 10.1242/jcs.252676] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
Membrane trafficking is an essential cellular process conserved across all eukaryotes, which regulates the uptake or release of macromolecules from cells, the composition of cellular membranes and organelle biogenesis. It influences numerous aspects of cellular organisation, dynamics and homeostasis, including nutrition, signalling and cell architecture. Not surprisingly, malfunction of membrane trafficking is linked to many serious genetic, metabolic and neurological disorders. It is also often hijacked during viral infection, enabling viruses to accomplish many of the main stages of their replication cycle, including entry into and egress from cells. The appropriation of membrane trafficking by viruses has been studied since the birth of cell biology and has helped elucidate how this integral cellular process functions. In this Review, we discuss some of the different strategies viruses use to manipulate and take over the membrane compartments of their hosts to promote their replication, assembly and egress.
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Affiliation(s)
- Miguel Hernandez-Gonzalez
- Cellular Signalling and Cytoskeletal Function Laboratory, The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK
| | - Gabrielle Larocque
- Cellular Signalling and Cytoskeletal Function Laboratory, The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK
| | - Michael Way
- Cellular Signalling and Cytoskeletal Function Laboratory, The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK
- Department of Infectious Disease, Imperial College, London W2 1PG, UK
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23
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Iossa D, Vitrone M, Gagliardi M, Falco E, Ragone E, Zampino R, Durante-Mangoni E. Anthropometric parameters and liver histology influence lipid metabolic changes in HCV chronic hepatitis on direct-acting antiviral treatment. ANNALS OF TRANSLATIONAL MEDICINE 2021; 9:35. [PMID: 33553328 PMCID: PMC7859777 DOI: 10.21037/atm-20-669] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Background Hepatitis C virus (HCV) infection affects lipid metabolism. We investigated the impact of direct-acting antiviral (DAA) treatment on lipid metabolism in chronic hepatitis C (CHC), with a focus on the effects of anthropometric parameters and liver histology. We also analyzed the dynamics of metabolic indexes used to estimate cardiovascular risk. Methods In 49 patients with CHC treated with DAAs, lipid metabolic changes, anthropometric parameters, liver histology and cardiovascular risk indexes, including triglyceride to HDL ratio (Tr/HDL), fatty liver index (FLI) and visceral adiposity index (VAI) were evaluated at baseline (BL), end of treatment (EOT) and 12 [sustained virological response (SVR) 12] and 24 (SVR24) weeks after EOT. Results SVR occurred in 96% of cases. Total and LDL cholesterol and ApoB levels increased significantly between BL and EOT (P<0.001, <0.001 and 0.05, respectively) and remained stable thereafter. Total and LDL cholesterol significantly increased only in patients with higher BL waist circumference (P<0.01 and 0.009), fibrosis (P=0.002 and 0.005) and steatosis (P=0.043 and 0.033, respectively). HDL cholesterol significantly rose at SVR24. However, cardiovascular risk indexes (Tr/HDL ratio, FLI and VAI) did not significantly change during DAA treatment and follow up. Conclusions Patients with HCV eradication after DAA treatment develop a pro-atherogenic lipid pattern, which varies according to anthropometric parameters and liver histology. However, no increase of cardiovascular risk indexes occurs in the short-term. Total and LDL cholesterol should be monitored long-term in CHC patients cured from infection.
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Affiliation(s)
- Domenico Iossa
- Internal Medicine, University of Campania "L. Vanvitelli", Naples, Italy
| | - Martina Vitrone
- Internal Medicine, University of Campania "L. Vanvitelli", Naples, Italy
| | - Massimo Gagliardi
- Internal Medicine, University of Campania "L. Vanvitelli", Naples, Italy
| | - Erasmo Falco
- Units of Infectious & Transplant Medicine AORN dei Colli, Monaldi Hospital, Naples, Italy
| | - Enrico Ragone
- Units of Infectious & Transplant Medicine AORN dei Colli, Monaldi Hospital, Naples, Italy
| | - Rosa Zampino
- Internal Medicine, University of Campania "L. Vanvitelli", Naples, Italy.,Units of Infectious & Transplant Medicine AORN dei Colli, Monaldi Hospital, Naples, Italy
| | - Emanuele Durante-Mangoni
- Internal Medicine, University of Campania "L. Vanvitelli", Naples, Italy.,Units of Infectious & Transplant Medicine AORN dei Colli, Monaldi Hospital, Naples, Italy
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24
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Zheng F, Li N, Xu Y, Zhou Y, Li YP. Adaptive mutations promote hepatitis C virus assembly by accelerating core translocation to the endoplasmic reticulum. J Biol Chem 2021; 296:100018. [PMID: 33144326 PMCID: PMC7949066 DOI: 10.1074/jbc.ra120.016010] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/10/2020] [Revised: 10/22/2020] [Accepted: 11/03/2020] [Indexed: 12/14/2022] Open
Abstract
The envelopment of hepatitis C virus (HCV) is believed to occur primarily in the endoplasmic reticulum (ER)-associated membrane, and the translocation of viral Core protein from lipid droplets (LDs) to the ER is essential for the envelopment of viral particles. However, the factors involved are not completely understood. Herein, we identified eight adaptive mutations that enhanced virus spread and infectivity of genotype 1a clone TNcc in hepatoma Huh7 cells through long-term culture adaptation and reverse genetic study. Of eight mutations, I853V in NS2 and C2865F in NS5B were found to be minimal mutation sets that enabled an increase in virus production without apparently affecting RNA replication, thus suggesting its roles in the post-replication stage of the HCV life cycle. Using a protease K protection and confocal microscopy analysis, we demonstrated that C2865F and the combination of I853V/C2865F enhanced virus envelopment by facilitating Core translocation from the LDs to the ER. Buoyant density analysis revealed that I853V/C2865F contributed to the release of virion with a density of ∼1.10 g/ml. Moreover, we demonstrated that NS5B directly interacted with NS2 at the protease domain and that mutations I853V, C2865F, and I853V/C2865F enhanced the interaction. In addition, C2865F also enhanced the interaction between NS5B and Core. In conclusion, this study demonstrated that adaptive mutations in NS2 and NS5B promoted HCV envelopment by accelerating Core translocation from the LDs to the ER and reinforced the interaction between NS2 and NS5B. The findings facilitate our understanding of the assembly of HCV morphogenesis.
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Affiliation(s)
- Fuxiang Zheng
- Institute of Human Virology, Zhongshan School of Medicine, and Key Laboratory of Tropical Disease Control of Ministry of Education, Sun Yat-sen University, Guangzhou, China
| | - Ni Li
- Institute of Human Virology, Zhongshan School of Medicine, and Key Laboratory of Tropical Disease Control of Ministry of Education, Sun Yat-sen University, Guangzhou, China
| | - Yi Xu
- Department of Pediatric, Guangzhou Women and Children's Medical Center, Guangzhou Medical University, Guangzhou, China
| | - Yuanping Zhou
- Department of Infectious Diseases, Nanfang Hospital, Southern Medical University, Guangzhou, China
| | - Yi-Ping Li
- Institute of Human Virology, Zhongshan School of Medicine, and Key Laboratory of Tropical Disease Control of Ministry of Education, Sun Yat-sen University, Guangzhou, China; Department of Infectious Disease, The Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China.
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25
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Shimotohno K. HCV Assembly and Egress via Modifications in Host Lipid Metabolic Systems. Cold Spring Harb Perspect Med 2021; 11:cshperspect.a036814. [PMID: 32122916 PMCID: PMC7778218 DOI: 10.1101/cshperspect.a036814] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/03/2023]
Abstract
Hepatitis C virus (HCV) proliferates by hijacking the host lipid machinery. In vitro replication systems revealed many aspects of the virus life cycle; in particular, viral utilization of host lipid metabolism during HCV proliferation. HCV interacts with lipid droplets (LDs) before starting the process of virus capsid formation at the lipid-rich endoplasmic reticulum (ER) membrane compartment. HCV buds into the ER via lipoprotein assembly and secretion. Exchangeable apolipoproteins, represented by apolipoprotein E (apoE), play pivotal roles in enhancing HCV-specific infectivity. HCV virions are likely to interact with other lipoproteins circulating in blood vessels and incorporate apolipoproteins as well as lipids. This review focuses on virus assembly and egress by briefly describing the recent advances in this area.
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26
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Alzahrani N, Wu MJ, Shanmugam S, Yi M. Delayed by Design: Role of Suboptimal Signal Peptidase Processing of Viral Structural Protein Precursors in Flaviviridae Virus Assembly. Viruses 2020; 12:v12101090. [PMID: 32993149 PMCID: PMC7601889 DOI: 10.3390/v12101090] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/29/2020] [Revised: 09/04/2020] [Accepted: 09/24/2020] [Indexed: 02/06/2023] Open
Abstract
The Flaviviridae virus family is classified into four different genera, including flavivirus, hepacivirus, pegivirus, and pestivirus, which cause significant morbidity and mortality in humans and other mammals, including ruminants and pigs. These are enveloped, single-stranded RNA viruses sharing a similar genome organization and replication scheme with certain unique features that differentiate them. All viruses in this family express a single polyprotein that encodes structural and nonstructural proteins at the N- and C-terminal regions, respectively. In general, the host signal peptidase cleaves the structural protein junction sites, while virus-encoded proteases process the nonstructural polyprotein region. It is known that signal peptidase processing is a rapid, co-translational event. Interestingly, certain signal peptidase processing site(s) in different Flaviviridae viral structural protein precursors display suboptimal cleavage kinetics. This review focuses on the recent progress regarding the Flaviviridae virus genus-specific mechanisms to downregulate signal peptidase-mediated processing at particular viral polyprotein junction sites and the role of delayed processing at these sites in infectious virus particle assembly.
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27
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Johri MK, Lashkari HV, Gupta D, Vedagiri D, Harshan KH. mTORC1 restricts hepatitis C virus RNA replication through ULK1-mediated suppression of miR-122 and facilitates post-replication events. J Gen Virol 2020; 101:86-95. [PMID: 31821132 DOI: 10.1099/jgv.0.001356] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/26/2022] Open
Abstract
The mechanistic target of rapamycin (mTOR), an important kinase that assimilates several upstream signals, associates into two functional complexes, mTORC1 and mTORC2. In this study, we demonstrate that HCV infection activates mTORC1 that functions in important antiviral response. Pharmacological inhibition of mTOR complexes augmented cellular HCV RNA levels, the observation confirmed further by Raptor depletion, indicating antiviral roles of mTORC1. ULK1 depletion phenocopied mTOR inhibition and thus suggested that mTORC1 restricts HCV replication through ULK1. We reveal that ULK1 depletion augmented the levels of miR-122, a critical host factor for HCV replication, thus possibly regulating HCV replication. The increase in HCV RNA levels, however, failed to augment intracellular infectious virion production, reflecting a lower rate of virion assembly. Higher intracellular HCV RNA levels, however, did not result in a corresponding increase in HCV RNA and infectious titres in mTOR inhibited supernatants, but in contrast showed a consistent drop, confirming defective viral assembly caused by the inhibition. Consistent with this, the mTOR activator caused a significant drop in HCV RNA levels both in infected cells and in the supernatant. Our results demonstrate that ULK1 depletion did not affect autophagy, suggesting that ULK1-mediated HCV regulation is autophagy independent. Together, our data demonstrate that mTORC1 functions to suppress HCV RNA replication, but facilitates the virion packaging and release. Our studies reveal that the activation of mTOR by HCV infection is an antiviral measure by the cells.
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Affiliation(s)
- Manish Kumar Johri
- Academy of Scientific and Innovative Research (AcSIR), CSIR-CCMB Campus, Hyderabad, India.,CSIR-Centre for Cellular and Molecular Biology, Uppal Road, Hyderabad-500007, India
| | | | - Divya Gupta
- CSIR-Centre for Cellular and Molecular Biology, Uppal Road, Hyderabad-500007, India
| | - Dhiviya Vedagiri
- Academy of Scientific and Innovative Research (AcSIR), CSIR-CCMB Campus, Hyderabad, India.,CSIR-Centre for Cellular and Molecular Biology, Uppal Road, Hyderabad-500007, India
| | - Krishnan Harinivas Harshan
- CSIR-Centre for Cellular and Molecular Biology, Uppal Road, Hyderabad-500007, India.,Academy of Scientific and Innovative Research (AcSIR), CSIR-CCMB Campus, Hyderabad, India
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28
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Lee JY, Cortese M, Haselmann U, Tabata K, Romero-Brey I, Funaya C, Schieber NL, Qiang Y, Bartenschlager M, Kallis S, Ritter C, Rohr K, Schwab Y, Ruggieri A, Bartenschlager R. Spatiotemporal Coupling of the Hepatitis C Virus Replication Cycle by Creating a Lipid Droplet- Proximal Membranous Replication Compartment. Cell Rep 2020; 27:3602-3617.e5. [PMID: 31216478 DOI: 10.1016/j.celrep.2019.05.063] [Citation(s) in RCA: 81] [Impact Index Per Article: 16.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/03/2019] [Revised: 04/05/2019] [Accepted: 05/17/2019] [Indexed: 02/08/2023] Open
Abstract
The hepatitis C virus (HCV) is a major cause of chronic liver disease, affecting around 71 million people worldwide. Viral RNA replication occurs in a membranous compartment composed of double-membrane vesicles (DMVs), whereas virus particles are thought to form by budding into the endoplasmic reticulum (ER). It is unknown how these steps are orchestrated in space and time. Here, we established an imaging system to visualize HCV structural and replicase proteins in live cells and with high resolution. We determined the conditions for the recruitment of viral proteins to putative assembly sites and studied the dynamics of this event and the underlying ultrastructure. Most notable was the selective recruitment of ER membranes around lipid droplets where structural proteins and the viral replicase colocalize. Moreover, ER membranes wrapping lipid droplets were decorated with double membrane vesicles, providing a topological map of how HCV might coordinate the steps of viral replication and virion assembly.
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Affiliation(s)
- Ji-Young Lee
- Department of Infectious Diseases, Molecular Virology, Heidelberg University, Im Neuenheimer Feld 344, 69120 Heidelberg, Germany; German Center for Infection Research, Heidelberg Partner Site, Im Neuenheimer Feld 344, 69120 Heidelberg, Germany
| | - Mirko Cortese
- Department of Infectious Diseases, Molecular Virology, Heidelberg University, Im Neuenheimer Feld 344, 69120 Heidelberg, Germany
| | - Uta Haselmann
- Department of Infectious Diseases, Molecular Virology, Heidelberg University, Im Neuenheimer Feld 344, 69120 Heidelberg, Germany
| | - Keisuke Tabata
- Department of Infectious Diseases, Molecular Virology, Heidelberg University, Im Neuenheimer Feld 344, 69120 Heidelberg, Germany
| | - Inés Romero-Brey
- Department of Infectious Diseases, Molecular Virology, Heidelberg University, Im Neuenheimer Feld 344, 69120 Heidelberg, Germany
| | - Charlotta Funaya
- Electron Microscopy Core Facility, Heidelberg University, 69120 Heidelberg, Germany
| | - Nicole L Schieber
- Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, 69117 Heidelberg, Germany
| | - Yu Qiang
- Biomedical Computer Vision Group, Heidelberg University, BIOQUANT, IPMB, and DKFZ Heidelberg, Im Neuenheimer Feld 267, 69120 Heidelberg, Germany
| | - Marie Bartenschlager
- Department of Infectious Diseases, Molecular Virology, Heidelberg University, Im Neuenheimer Feld 344, 69120 Heidelberg, Germany
| | - Stephanie Kallis
- Department of Infectious Diseases, Molecular Virology, Heidelberg University, Im Neuenheimer Feld 344, 69120 Heidelberg, Germany
| | - Christian Ritter
- Biomedical Computer Vision Group, Heidelberg University, BIOQUANT, IPMB, and DKFZ Heidelberg, Im Neuenheimer Feld 267, 69120 Heidelberg, Germany
| | - Karl Rohr
- Biomedical Computer Vision Group, Heidelberg University, BIOQUANT, IPMB, and DKFZ Heidelberg, Im Neuenheimer Feld 267, 69120 Heidelberg, Germany
| | - Yannick Schwab
- Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, 69117 Heidelberg, Germany; Electron Microscopy Core Facility, European Molecular Biology Laboratory, 69117 Heidelberg, Germany
| | - Alessia Ruggieri
- Department of Infectious Diseases, Molecular Virology, Heidelberg University, Im Neuenheimer Feld 344, 69120 Heidelberg, Germany
| | - Ralf Bartenschlager
- Department of Infectious Diseases, Molecular Virology, Heidelberg University, Im Neuenheimer Feld 344, 69120 Heidelberg, Germany; German Center for Infection Research, Heidelberg Partner Site, Im Neuenheimer Feld 344, 69120 Heidelberg, Germany; Division of Virus-Associated Carcinogenesis, German Cancer Research Center (DKFZ), Im Neuenheimer Feld 242, Heidelberg, Germany.
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29
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Martínez JL, Arias CF. Role of the Guanine Nucleotide Exchange Factor GBF1 in the Replication of RNA Viruses. Viruses 2020; 12:E682. [PMID: 32599855 PMCID: PMC7354614 DOI: 10.3390/v12060682] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/10/2020] [Revised: 04/06/2020] [Accepted: 04/06/2020] [Indexed: 12/12/2022] Open
Abstract
The guanine nucleotide exchange factor GBF1 is a well-known factor that can activate different ADP-ribosylation factor (Arf) proteins during the regulation of different cellular vesicular transport processes. In the last decade, it has become increasingly evident that GBF1 can also regulate different steps of the replication cycle of RNA viruses belonging to different virus families. GBF1 has been shown not only to facilitate the intracellular traffic of different viral and cellular elements during infection, but also to modulate the replication of viral RNA, the formation and maturation of viral replication complexes, and the processing of viral proteins through mechanisms that do not depend on its canonical role in intracellular transport. Here, we review the various roles that GBF1 plays during the replication of different RNA viruses.
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Affiliation(s)
| | - Carlos F. Arias
- Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca 4510, Morelos, Mexico;
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30
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Cosset FL, Mialon C, Boson B, Granier C, Denolly S. HCV Interplay with Lipoproteins: Inside or Outside the Cells? Viruses 2020; 12:v12040434. [PMID: 32290553 PMCID: PMC7232430 DOI: 10.3390/v12040434] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2020] [Revised: 04/05/2020] [Accepted: 04/10/2020] [Indexed: 12/11/2022] Open
Abstract
Hepatitis C virus (HCV) infection is a major public health issue leading to chronic liver diseases. HCV particles are unique owing to their particular lipid composition, namely the incorporation of neutral lipids and apolipoproteins. The mechanism of association between HCV virion components and these lipoproteins factors remains poorly understood as well as its impact in subsequent steps of the viral life cycle, such as entry into cells. It was proposed that the lipoprotein biogenesis pathway is involved in HCV morphogenesis; yet, recent evidence indicated that HCV particles can mature and evolve biochemically in the extracellular medium after egress. In addition, several viral, cellular and blood components have been shown to influence and regulate this specific association. Finally, this specific structure and composition of HCV particles was found to influence entry into cells as well as their stability and sensitivity to neutralizing antibodies. Due to its specific particle composition, studying the association of HCV particles with lipoproteins remains an important goal towards the rational design of a protective vaccine.
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31
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Cai H, Yao W, Huang J, Xiao J, Chen W, Hu L, Mai R, Liang M, Chen D, Jiang N, Zhou L, Peng T. Apolipoprotein M, identified as a novel hepatitis C virus (HCV) particle associated protein, contributes to HCV assembly and interacts with E2 protein. Antiviral Res 2020; 177:104756. [PMID: 32119870 DOI: 10.1016/j.antiviral.2020.104756] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/19/2019] [Revised: 01/18/2020] [Accepted: 02/25/2020] [Indexed: 02/08/2023]
Abstract
Hepatitis C virus (HCV) infection is a major cause of chronic liver diseases such as steatosis, cirrhosis, and hepatocellular carcinoma. HCV particles have been found to associate with apolipoproteins, and apolipoproteins not only participate in the HCV life cycle, but also help HCV escape recognition by the host immune system, which pose challenges for the development of both HCV treatments and vaccines. However, no study has reported on the comprehensive identification of apolipoprotein associations with HCV particles. In the present study, we performed proteome analysis by affinity purification coupled with mass spectrometry (AP-MS) to comprehensively identify the apolipoprotein associations with HCV particles, and ApoM was first identified by AP-MS besides the previously reported ApoE, ApoB, ApoA-I and ApoC-I. Additionally, three assays further confirmed that ApoM was a novel virus particle associated protein. We also showed that ApoM was required for HCV production, especially for the assembly/release step of HCV life cycle. Furthermore, ApoM interacted with the HCV E2 protein. Finally, HCV infection reduced ApoM expression both in vitro and in vivo. Collectively, our study demonstrates that ApoM, identified as a novel HCV particle associated protein, contributes to HCV assembly/release and interacts with HCV E2 protein. It provides new insights on how HCV and the host apolipoproteins are reciprocally influenced and lays a basis for research in developing innovative antiviral strategies.
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Affiliation(s)
- Hua Cai
- Guangzhou Hoffmann Institute of Immunology, College of Basic Sciences, The Fifth Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University, Guangzhou, China
| | - Wenxia Yao
- Guangzhou Hoffmann Institute of Immunology, College of Basic Sciences, The Fifth Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University, Guangzhou, China.
| | - Jingxian Huang
- Guangzhou Hoffmann Institute of Immunology, College of Basic Sciences, The Fifth Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University, Guangzhou, China
| | - Jing Xiao
- Guangzhou Hoffmann Institute of Immunology, College of Basic Sciences, The Fifth Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University, Guangzhou, China
| | - Wenli Chen
- Department of Infectious Diseases, Guangdong Provincial People's Hospital, Guangzhou, China
| | - Longbo Hu
- Guangzhou Hoffmann Institute of Immunology, College of Basic Sciences, The Fifth Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University, Guangzhou, China
| | - Runming Mai
- Guangzhou Hoffmann Institute of Immunology, College of Basic Sciences, The Fifth Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University, Guangzhou, China
| | - Mengdi Liang
- Guangzhou Hoffmann Institute of Immunology, College of Basic Sciences, The Fifth Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University, Guangzhou, China
| | - Di Chen
- Guangzhou Hoffmann Institute of Immunology, College of Basic Sciences, The Fifth Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University, Guangzhou, China
| | - Nan Jiang
- The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China
| | - Li Zhou
- The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou, China
| | - Tao Peng
- Guangzhou Hoffmann Institute of Immunology, College of Basic Sciences, The Fifth Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University, Guangzhou, China.
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32
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Li X, Li J, Feng Y, Cai H, Li YP, Peng T. Long-chain fatty acyl-coenzyme A suppresses hepatitis C virus infection by targeting virion-bound lipoproteins. Antiviral Res 2020; 177:104734. [PMID: 32057770 DOI: 10.1016/j.antiviral.2020.104734] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/26/2019] [Revised: 12/18/2019] [Accepted: 02/03/2020] [Indexed: 12/12/2022]
Abstract
Hepatitis C virus (HCV) is a leading cause of chronic hepatitis and end-stage liver diseases. Mature HCV virions are bound by host-derived lipoproteins. Lack of an HCV vaccine warrants a major role of antiviral treatment in the global elimination of hepatitis C. Although direct-acting antivirals (DAAs) are replacing the interferon-based treatment and have dramatically improved the cure rate, the presence of viral variants resistant to DAAs, HCV genotype/subtype-specific efficacy, and high cost of DAAs argue novel and affordable regimens. In this study, we identified the antiviral effects of long-chain fatty acyl-coenzyme A (LCFA-CoA) against the infections of HCV genotypes 1-6 through targeting mature HCV-bound lipoproteins, suggesting novel mechanism(s) of antiviral different from those used by host-targeting agents or DAAs. We found that the antiviral activity of LCFA-CoA relied on the long-chain saturated fatty acid and the CoA group, and was enhanced when combined with pegylated-interferon or DAAs. Importantly, we demonstrated that LCFA-CoA efficiently inhibited the infection of HCV variants carrying DAA-resistant mutations. The mechanistic study revealed that LCFA-CoA specifically abolished the attachment and binding steps and also inhibited the cell-to-cell viral transmission. LCFA-CoA targeted mature HCV-bound lipoproteins, but not apolipoproteins B or E. In addition, LCFA-CoA could also inhibit the infection of the dengue virus. Our findings suggest that LCFA-CoA could potentially serve as a supplement HCV therapy, particularly for the DAA-resistant HCV variants. Taken together, LCFA-CoA may be further developed to be a novel class of antivirals with mechanism(s), different from host-targeting agents or DAAs, of targeting the components associated with mature HCV virions.
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Affiliation(s)
- Xinlei Li
- State Key Laboratory of Respiratory Disease, Sino-French Hoffmann Institute, Guangzhou Medical University, Guangzhou, China
| | - Jinqian Li
- Institute of Human Virology, Zhongshan School of Medicine, Key Laboratory of Tropical Disease Control (Sun Yat-sen University), Ministry of Education, Sun Yat-sen University, Guangzhou, 510080, China
| | - Yetong Feng
- Institute of Human Virology, Zhongshan School of Medicine, Key Laboratory of Tropical Disease Control (Sun Yat-sen University), Ministry of Education, Sun Yat-sen University, Guangzhou, 510080, China
| | - Hua Cai
- State Key Laboratory of Respiratory Disease, Sino-French Hoffmann Institute, Guangzhou Medical University, Guangzhou, China
| | - Yi-Ping Li
- Institute of Human Virology, Zhongshan School of Medicine, Key Laboratory of Tropical Disease Control (Sun Yat-sen University), Ministry of Education, Sun Yat-sen University, Guangzhou, 510080, China.
| | - Tao Peng
- State Key Laboratory of Respiratory Disease, Sino-French Hoffmann Institute, Guangzhou Medical University, Guangzhou, China.
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33
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The Host Factor Erlin-1 is Required for Efficient Hepatitis C Virus Infection. Cells 2019; 8:cells8121555. [PMID: 31810281 PMCID: PMC6953030 DOI: 10.3390/cells8121555] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/28/2019] [Revised: 11/28/2019] [Accepted: 11/29/2019] [Indexed: 12/22/2022] Open
Abstract
Development of hepatitis C virus (HCV) infection cell culture systems has permitted the identification of cellular factors that regulate the HCV life cycle. Some of these cellular factors affect steps in the viral life cycle that are tightly associated with intracellular membranes derived from the endoplasmic reticulum (ER). Here, we describe the discovery of erlin-1 protein as a cellular factor that regulates HCV infection. Erlin-1 is a cholesterol-binding protein located in detergent-resistant membranes within the ER. It is implicated in cholesterol homeostasis and the ER-associated degradation pathway. Silencing of erlin-1 protein expression by siRNA led to decreased infection efficiency characterized by reduction in intracellular RNA accumulation, HCV protein expression and virus production. Mechanistic studies revealed that erlin-1 protein is required early in the infection, downstream of cell entry and primary translation, specifically to initiate RNA replication, and later in the infection to support infectious virus production. This study identifies erlin-1 protein as an important cellular factor regulating HCV infection.
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34
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Castro V, Calvo G, Ávila-Pérez G, Dreux M, Gastaminza P. Differential Roles of Lipin1 and Lipin2 in the Hepatitis C Virus Replication Cycle. Cells 2019; 8:cells8111456. [PMID: 31752156 PMCID: PMC6912735 DOI: 10.3390/cells8111456] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/18/2019] [Revised: 11/09/2019] [Accepted: 11/13/2019] [Indexed: 02/06/2023] Open
Abstract
Although their origin, nature and structure are not identical, a common feature of positive-strand RNA viruses is their ability to subvert host lipids and intracellular membranes to generate replication and assembly complexes. Recently, lipin1, a cellular enzyme that converts phosphatidic acid into diacylglycerol, has been implicated in the formation of the membranous web that hosts hepatitis C virus (HCV) replicase. In the liver, lipin1 cooperates with lipin2 to maintain glycerolipid homeostasis. We extended our previous study of the lipin family on HCV infection, by determining the impact of the lipin2 silencing on viral replication. Our data reveal that lipin2 silencing interferes with HCV virion secretion at late stages of the infection, without significantly affecting viral replication or assembly. Moreover, uninfected lipin2-, but not lipin1-deficient cells display alterations in mitochondrial and Golgi apparatus morphology, suggesting that lipin2 contributes to the maintenance of the overall organelle architecture. Finally, our data suggest a broader function of lipin2 for replication of HCV and other RNA viruses, in contrast with the specific impact of lipin1 silencing on HCV replication. Overall, this study reveals distinctive functions of lipin1 and lipin2 in cells of hepatic origin, a context in which they are often considered functionally redundant.
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Affiliation(s)
- Victoria Castro
- Department of Cellular and Molecular Biology Centro Nacional de Biotecnología-Consejo Superior de Investigaciones Científicas, Centro Nacional de Biotecnología-C.S.I.C., Calle Darwin 3, 28049 Madrid, Spain; (V.C.); (G.C.); (G.Á.-P.)
| | - Gema Calvo
- Department of Cellular and Molecular Biology Centro Nacional de Biotecnología-Consejo Superior de Investigaciones Científicas, Centro Nacional de Biotecnología-C.S.I.C., Calle Darwin 3, 28049 Madrid, Spain; (V.C.); (G.C.); (G.Á.-P.)
| | - Ginés Ávila-Pérez
- Department of Cellular and Molecular Biology Centro Nacional de Biotecnología-Consejo Superior de Investigaciones Científicas, Centro Nacional de Biotecnología-C.S.I.C., Calle Darwin 3, 28049 Madrid, Spain; (V.C.); (G.C.); (G.Á.-P.)
| | - Marlène Dreux
- CIRI, Inserm, U1111, Université Claude Bernard Lyon 1, CNRS, UMR5308, École Normale Supérieure de Lyon, Univ Lyon, F-69007 Lyon, France;
| | - Pablo Gastaminza
- Department of Cellular and Molecular Biology Centro Nacional de Biotecnología-Consejo Superior de Investigaciones Científicas, Centro Nacional de Biotecnología-C.S.I.C., Calle Darwin 3, 28049 Madrid, Spain; (V.C.); (G.C.); (G.Á.-P.)
- Correspondence: ; Tel.: +34-91-585-4678; Fax: +34-91-585-4506
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Abouelasrar Salama S, Lavie M, De Buck M, Van Damme J, Struyf S. Cytokines and serum amyloid A in the pathogenesis of hepatitis C virus infection. Cytokine Growth Factor Rev 2019; 50:29-42. [PMID: 31718982 DOI: 10.1016/j.cytogfr.2019.10.006] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/29/2019] [Revised: 10/17/2019] [Accepted: 10/21/2019] [Indexed: 02/07/2023]
Abstract
Expression of the acute phase protein serum amyloid A (SAA) is dependent on the release of the pro-inflammatory cytokines IL-1, IL-6 and TNF-α during infection and inflammation. Hepatitis C virus (HCV) upregulates SAA-inducing cytokines. In line with this, a segment of chronically infected individuals display increased circulating levels of SAA. SAA has even been proposed to be a potential biomarker to evaluate treatment efficiency and the course of disease. SAA possesses antiviral activity against HCV via direct interaction with the viral particle, but might also divert infectivity through its function as an apolipoprotein. On the other hand, SAA shares inflammatory and angiogenic activity with chemotactic cytokines by activating the G protein-coupled receptor, formyl peptide receptor 2. These latter properties might promote chronic inflammation and hepatic injury. Indeed, up to 80 % of infected individuals develop chronic disease because they cannot completely clear the infection, due to diversion of the immune response. In this review, we summarize the interconnection between SAA and cytokines in the context of HCV infection and highlight the dual role SAA could play in this disease. Nevertheless, more research is needed to establish whether the balance between those opposing activities can be tilted in favor of the host defense.
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Affiliation(s)
- Sara Abouelasrar Salama
- Laboratory of Molecular Immunology, Department of Microbiology, Immunology and Transplantation, Rega Institute for Medical Research, University of Leuven, Leuven, 3000, Belgium
| | - Muriel Lavie
- University of Lille, CNRS, Inserm, CHU Lille, Institut Pasteur de Lille, U1019, UMR 8204, Centre d'Infection et d'Immunité de Lille, Lille, France
| | - Mieke De Buck
- Laboratory of Molecular Immunology, Department of Microbiology, Immunology and Transplantation, Rega Institute for Medical Research, University of Leuven, Leuven, 3000, Belgium
| | - Jo Van Damme
- Laboratory of Molecular Immunology, Department of Microbiology, Immunology and Transplantation, Rega Institute for Medical Research, University of Leuven, Leuven, 3000, Belgium
| | - Sofie Struyf
- Laboratory of Molecular Immunology, Department of Microbiology, Immunology and Transplantation, Rega Institute for Medical Research, University of Leuven, Leuven, 3000, Belgium.
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Fukuhara T, Matsuura Y. Roles of secretory glycoproteins in particle formation of Flaviviridae viruses. Microbiol Immunol 2019; 63:401-406. [PMID: 31342548 DOI: 10.1111/1348-0421.12733] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/25/2019] [Revised: 07/17/2019] [Accepted: 07/18/2019] [Indexed: 12/12/2022]
Abstract
The family Flaviviridae comprises four genera, namely, Flavivirus, Pestivirus, Pegivirus, and Hepacivirus. These viruses have similar genome structures, but the genomes of Pestivirus and Flavivirus encode the secretory glycoproteins Erns and NS1, respectively. Erns plays an important role in virus particle formation and cell entry, whereas NS1 participates in the formation of replication complexes and virus particles. Conversely, apolipoproteins are known to participate in the formation of infectious particles of hepatitis C virus (HCV) and various secretory glycoproteins play a similar role in HCV particles formation, suggesting that there is no strong specificity for the function of secretory glycoproteins in infectious-particle formation. In addition, recent studies have shown that host-derived apolipoproteins and virus-derived Erns and NS1 play comparable roles in infectious-particle formation of both HCV and pestiviruses. In this review, we summarize the roles of secretory glycoproteins in the formation of Flaviviridae virus particles.
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Affiliation(s)
- Takasuke Fukuhara
- Department of Molecular Virology, Research Institute for Microbial Diseases, Osaka University, Osaka, Japan
| | - Yoshiharu Matsuura
- Department of Molecular Virology, Research Institute for Microbial Diseases, Osaka University, Osaka, Japan
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37
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Qiao L, Luo GG. Human apolipoprotein E promotes hepatitis B virus infection and production. PLoS Pathog 2019; 15:e1007874. [PMID: 31393946 PMCID: PMC6687101 DOI: 10.1371/journal.ppat.1007874] [Citation(s) in RCA: 32] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/20/2019] [Accepted: 05/27/2019] [Indexed: 12/16/2022] Open
Abstract
Hepatitis B virus (HBV) is a common cause of liver diseases, including chronic hepatitis, steatosis, fibrosis, cirrhosis, and hepatocellular carcinoma (HCC). HBV chronically infects about 240 million people worldwide, posing a major global health problem. The current standard antiviral therapy effectively inhibits HBV replication but does not eliminate the virus unlike direct-acting antivirals (DAA) for curing hepatitis C. Our previous studies have demonstrated that human apolipoprotein E (apoE) plays important roles in hepatitis C virus infection and morphogenesis. In the present study, we have found that apoE is also associated with HBV and is required for efficient HBV infection. An apoE-specific monoclonal antibody was able to capture HBV similar to anti-HBs. More importantly, apoE monoclonal antibody could effectively block HBV infection, resulting in a greater than 90% reduction of HBV infectivity. Likewise, silencing of apoE expression or knockout of apoE gene by CRISPR/Cas9 resulted in a greater than 90% reduction of HBV infection and more than 80% decrease of HBV production, which could be fully restored by ectopic apoE expression. However, apoE silencing or knockout did not significantly affect HBV DNA replication or the production of nonenveloped (naked) nucleocapsids. These findings demonstrate that human apoE promotes HBV infection and production. We speculate that apoE may also play a role in persistent HBV infection by evading host immune response similar to its role in the HCV life cycle and pathogenesis. Inhibitors interfering with apoE biogenesis, secretion, and/or binding to receptors may serve as antivirals for elimination of chronic HBV infection.
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Affiliation(s)
- Luhua Qiao
- Department of Microbiology, University of Alabama at Birmingham School of Medicine, Birmingham, Alabama, United States of America
| | - Guangxiang George Luo
- Department of Microbiology, University of Alabama at Birmingham School of Medicine, Birmingham, Alabama, United States of America
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38
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[Hepatic tropism of hepatitis C virus infection]. Uirusu 2019; 68:63-70. [PMID: 31105136 DOI: 10.2222/jsv.68.63] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/26/2022]
Abstract
Hepatitis C virus (HCV) infects over 170 million people worldwide and is a major cause of life-threatening liver diseases such as liver cirrhosis and hepatocellular carcinoma. In current research, we aimed to clarify the mechanism of hepatic tropism of HCV infection. Although non-hepatic cells could not permit replication of HCV RNA, exogenous expression of liver specific miRNA, miR-122 facilitated efficient replication of viral RNA through direct interaction with 5'UTR of viral genome, indicating that miR-122 is one of the key determinants for hepatic tropism of HCV infection. In spite of efficient replication of viral RNA, formation of infectious particles was not observed in non-hepatic cells exogenously expressing miR-122. We found that expression of apolipoprotein E (ApoE) facilitated the formation of infectious HCV particles in non-hepatic cells, indicating that not only miR-122 but also ApoE participate in tissue tropism of HCV infection. To understand the exact roles of miR-122 and apolipoproteins in hepatic tropism of HCV, we established miR-122 and ApoB/ApoE knockout (KO) Huh7 cells, respectively. Although slight increase of intracellular HCV RNA and infectious titers in the culture supernatants was observed, propagation of HCV was impaired in miR-122 KO Huh7 cells. After serial passages of HCV in miR-122 KO cells, we obtained an adaptive mutant that possessed G28A substitutions in the 5'UTR of the HCV genome and exhibited efficient translation and replication in both miR-122 KO Huh7 and non-hepatic cells without exogenous expression of miR-122. These results suggest that HCV mutants replicating in non-hepatic cells in an miR-122-independent manner participate in the induction of extrahepatic manifestations in chronic hepatitis C patients. Deficiency of both ApoB and ApoE strongly inhibited the formation of infectious HCV particles. Interestingly, expression not only of ApoE but also of ApoA or ApoC could rescue the production of infectious HCV particles in ApoB/ApoE KO cells, suggesting that exchangeable apolipoproteins redundantly participate in the formation of infectious HCV particles.
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39
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Collett S, Torresi J, Earnest-Silveira L, Christiansen D, Elbourne A, Ramsland PA. Probing and pressing surfaces of hepatitis C virus-like particles. J Colloid Interface Sci 2019; 545:259-268. [DOI: 10.1016/j.jcis.2019.03.022] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/14/2019] [Revised: 03/07/2019] [Accepted: 03/09/2019] [Indexed: 02/09/2023]
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40
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Vieyres G, Pietschmann T. HCV Pit Stop at the Lipid Droplet: Refuel Lipids and Put on a Lipoprotein Coat before Exit. Cells 2019; 8:cells8030233. [PMID: 30871009 PMCID: PMC6468556 DOI: 10.3390/cells8030233] [Citation(s) in RCA: 41] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/28/2019] [Revised: 02/28/2019] [Accepted: 03/04/2019] [Indexed: 02/07/2023] Open
Abstract
The replication cycle of the liver-tropic hepatitis C virus (HCV) is tightly connected to the host lipid metabolism, during the virus entry, replication, assembly and egress stages, but also while the virus circulates in the bloodstream. This interplay coins viral particle properties, governs viral cell tropism, and facilitates immune evasion. This review summarizes our knowledge of these interactions focusing on the late steps of the virus replication cycle. It builds on our understanding of the cell biology of lipid droplets and the biosynthesis of liver lipoproteins and attempts to explain how HCV hijacks these organelles and pathways to assemble its lipo-viro-particles. In particular, this review describes (i) the mechanisms of viral protein translocation to and from the lipid droplet surface and the orchestration of an interface between replication and assembly complexes, (ii) the importance of the triglyceride mobilization from the lipid droplets for HCV assembly, (iii) the interplay between HCV and the lipoprotein synthesis pathway including the role played by apolipoproteins in virion assembly, and finally (iv) the consequences of these complex virus–host interactions on the virion composition and its biophysical properties. The wealth of data accumulated in the past years on the role of the lipid metabolism in HCV assembly and its imprint on the virion properties will guide vaccine design efforts and reinforce our understanding of the hepatic lipid metabolism in health and disease.
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Affiliation(s)
- Gabrielle Vieyres
- Institute of Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a Joint Venture between the Medical School Hannover (MHH) and the Helmholtz Centre for Infection Research (HZI), 30625 Hannover, Germany.
| | - Thomas Pietschmann
- Institute of Experimental Virology, TWINCORE, Centre for Experimental and Clinical Infection Research, a Joint Venture between the Medical School Hannover (MHH) and the Helmholtz Centre for Infection Research (HZI), 30625 Hannover, Germany.
- German Centre for Infection Research (DZIF), Partner Site Hannover-Braunschweig, 38124 Braunschweig, Germany.
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41
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Roder AE, Vazquez C, Horner SM. The acidic domain of the hepatitis C virus NS4A protein is required for viral assembly and envelopment through interactions with the viral E1 glycoprotein. PLoS Pathog 2019; 15:e1007163. [PMID: 30730994 PMCID: PMC6382253 DOI: 10.1371/journal.ppat.1007163] [Citation(s) in RCA: 6] [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: 06/14/2018] [Revised: 02/20/2019] [Accepted: 01/05/2019] [Indexed: 02/07/2023] Open
Abstract
Hepatitis C virus (HCV) assembly and envelopment are coordinated by a complex protein interaction network that includes most of the viral structural and nonstructural proteins. While the nonstructural protein 4A (NS4A) is known to be important for viral particle production, the specific function of NS4A in this process is not well understood. We performed mutagenesis of the C-terminal acidic domain of NS4A and found that mutation of several of these amino acids prevented the formation of the viral envelope, and therefore the production of infectious virions, without affecting viral RNA replication. In an overexpression system, we found that NS4A interacted with several viral proteins known to coordinate envelopment, including the viral E1 glycoprotein. One of the NS4A C-terminal mutations, Y45F, disrupted the interaction of NS4A with E1. Specifically, NS4A interacted with the first hydrophobic region of E1, a region previously described as regulating viral particle production. Indeed, we found that an E1 mutation in this region, D72A, also disrupted the interaction of NS4A with E1. Supernatants from HCV NS4A Y45F transfected cells had significantly reduced levels of HCV RNA, however they contained equivalent levels of Core protein. Interestingly, the Core protein secreted from these cells formed high order oligomers with a density matching the infectious virus secreted from wild-type cells. These results suggest that this Y45F mutation in NS4A causes secretion of low-density Core particles lacking genomic HCV RNA. These results corroborate previous findings showing that the E1 D72A mutation also causes secretion of Core complexes lacking genomic HCV RNA, and therefore suggest that the interaction between NS4A and E1 is involved in the incorporation of viral RNA into infectious HCV particles. Our findings define a new role for NS4A in the HCV lifecycle and help elucidate the protein interactions necessary for production of infectious virus.
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Affiliation(s)
- Allison E Roder
- Department of Molecular Genetics & Microbiology, Duke University Medical Center, Durham, NC, United States of America
| | - Christine Vazquez
- Department of Molecular Genetics & Microbiology, Duke University Medical Center, Durham, NC, United States of America
| | - Stacy M Horner
- Department of Molecular Genetics & Microbiology, Duke University Medical Center, Durham, NC, United States of America
- Department of Medicine, Duke University Medical Center, Durham, NC, United States of America
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42
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Lassen S, Grüttner C, Nguyen-Dinh V, Herker E. Perilipin-2 is critical for efficient lipoprotein and hepatitis C virus particle production. J Cell Sci 2019; 132:jcs.217042. [PMID: 30559250 DOI: 10.1242/jcs.217042] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/20/2018] [Accepted: 12/07/2018] [Indexed: 12/18/2022] Open
Abstract
In hepatocytes, PLIN2 is the major protein coating lipid droplets (LDs), an organelle the hepatitis C virus (HCV) hijacks for virion morphogenesis. We investigated the consequences of PLIN2 deficiency on LDs and on HCV infection. Knockdown of PLIN2 did not affect LD homeostasis, likely due to compensation by PLIN3, but severely impaired HCV particle production. PLIN2-knockdown cells had slightly larger LDs with altered protein composition, enhanced local lipase activity and higher β-oxidation capacity. Electron micrographs showed that, after PLIN2 knockdown, LDs and HCV-induced vesicular structures were tightly surrounded by ER-derived double-membrane sacs. Strikingly, the LD access for HCV core and NS5A proteins was restricted in PLIN2-deficient cells, which correlated with reduced formation of intracellular HCV particles that were less infectious and of higher density, indicating defects in maturation. PLIN2 depletion also reduced protein levels and secretion of ApoE due to lysosomal degradation, but did not affect the density of ApoE-containing lipoproteins. However, ApoE overexpression in PLIN2-deficient cells did not restore HCV spreading. Thus, PLIN2 expression is required for trafficking of core and NS5A proteins to LDs, and for formation of functional low-density HCV particles prior to ApoE incorporation.This article has an associated First Person interview with the first author of the paper.
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Affiliation(s)
- Susan Lassen
- Heinrich Pette Institute, Leibniz Institute for Experimental Virology, 20251 Hamburg, Germany
| | - Cordula Grüttner
- Heinrich Pette Institute, Leibniz Institute for Experimental Virology, 20251 Hamburg, Germany
| | - Van Nguyen-Dinh
- Heinrich Pette Institute, Leibniz Institute for Experimental Virology, 20251 Hamburg, Germany
| | - Eva Herker
- Heinrich Pette Institute, Leibniz Institute for Experimental Virology, 20251 Hamburg, Germany .,Institute of Virology, Philipps University Marburg, 35043 Marburg, Germany
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Abstract
Apolipoprotein E (apoE) plays dual functions in the HCV life cycle by promoting HCV infection and virion assembly and production. ApoE is a structural component on the HCV envelope. It mediates HCV cell attachment through specific interactions with the cell surface receptors such as syndecan-1 (SDC-1) and SDC-2 heparan sulfate proteoglycans (HSPGs). It also interacts with NS5A and E2, resulting in an enhancement of HCV morphogenesis. It can bind HCV extracellularly and promotes HCV infection. It is critical for HCV cell-to-cell transmission and may also play a role in HCV persistence by interfering with the action of HCV-neutralizing antibodies. Other apolipoproteins particularly apoB and apoC1 were also found on the HCV envelope, but their roles in the HCV life cycle remain unclear. In the last decade, a number of genomic, immunological, structural, and cell biology methodologies have been developed and used for determining the importance of apoE in the HCV life cycle. These methods and protocols will continue to be valuable to further understand the importance and the underlying molecular mechanism of various apolipoproteins in HCV infection and pathogenesis.
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Affiliation(s)
- Luhua Qiao
- Department of Microbiology, University of Alabama at Birmingham School of Medicine, Birmingham, AL, USA
| | - Guangxiang George Luo
- Department of Microbiology, University of Alabama at Birmingham School of Medicine, Birmingham, AL, USA.
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44
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Castro V, Ávila-Pérez G, Mingorance L, Gastaminza P. A Cell Culture Model for Persistent HCV Infection. Methods Mol Biol 2019; 1911:157-168. [PMID: 30593624 DOI: 10.1007/978-1-4939-8976-8_10] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
Chronic hepatitis C virus (HCV) infection affects millions of humans throughout the globe, causing liver disease and hepatocellular carcinoma. Persistence of the virus in the infected host can last for decades as a result of a faulty immune response that fails to clear the virus while constituting a major player in viral pathogenesis. In addition to evading immune responses, HCV has evolved intracellular survival strategies that enable persistent replication without directly killing the host cell.After the generation of cell culture infection models for HCV, the knowledge about this virus and host-virus interactions acquired in the last decade has been greatly increased. Interestingly, persistent infection can also be established in cell culture. This model recapitulates persistent HCV RNA replication and viral protein expression as well as infectious progeny virus assembly and secretion and may be used to study not only these aspects of the virus replication cycle but also to study host-virus interactions in a model of prolonged HCV infection. In this chapter, we describe a methodology to generate persistently HCV-infected cultures and to monitor viral load and progeny virus production. Also, we provide generic protocols to study the impact of chemical compounds and host-targeting shRNAs to illustrate the applications of this model in the study of HCV infection in cell culture.
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Affiliation(s)
- Victoria Castro
- Department of Molecular and Cellular Biology, Centro Nacional De Biotecnología-Consejo Superior de Investigaciones Científicas (CNB-CSIC), Madrid, Spain
| | - Ginés Ávila-Pérez
- Department of Molecular and Cellular Biology, Centro Nacional De Biotecnología-Consejo Superior de Investigaciones Científicas (CNB-CSIC), Madrid, Spain
| | - Lidia Mingorance
- Department of Molecular and Cellular Biology, Centro Nacional De Biotecnología-Consejo Superior de Investigaciones Científicas (CNB-CSIC), Madrid, Spain
| | - Pablo Gastaminza
- Department of Molecular and Cellular Biology, Centro Nacional De Biotecnología-Consejo Superior de Investigaciones Científicas (CNB-CSIC), Madrid, Spain.
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45
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Park SB, Boyer A, Hu Z, Le D, Liang TJ. Discovery and characterization of a novel HCV inhibitor targeting the late stage of HCV life cycle. Antivir Ther 2019; 24:371-381. [PMID: 30880685 PMCID: PMC11542171 DOI: 10.3851/imp3303] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 03/03/2019] [Indexed: 10/27/2022]
Abstract
BACKGROUND Currently approved anti-HCV drugs, the direct-acting antivirals (DAAs), are highly effective and target the viral RNA replication stage of the HCV life cycle. Due to high mutation rate of HCV, drug resistant variants can arise during DAA monotherapy. Thus, a combination of DAAs is necessary to achieve a high response rate. Novel HCV inhibitors targeting the HCV late stage such as assembly and release may further improve combination therapy with the DAAs. Here we characterize one late stage-targeting candidate compound, 6-(4-chloro-3-methylphenoxy)-pyridin-3-amine (MLS000833705). METHODS We treated HCV-infected cells with MLS000833705 and other HCV inhibitors and examined HCV RNA and infectious titres. We evaluated the colocalization of HCV core and lipid droplets by confocal microscopy. We performed HCV core-proteinase K digestion assay and several lipid assays to study the mechanism of MLS000833705. RESULTS We showed that MLS000833705 decreased extracellular HCV RNA levels more than intracellular HCV RNA levels in HCV infectious cell culture. Similarly, MLS000833705 reduced infectious HCV titres substantially more in the culture supernatant than intracellularly. Confocal microscopy showed that MLS000833705 did not affect the colocalization of HCV core protein with cellular lipid droplets where HCV assembles. HCV core-proteinase K digestion assay showed that MLS000833705 inhibited the envelopment of HCV capsid. CONCLUSIONS Our study demonstrates that MLS000833705 is a late-stage HCV inhibitor targeting HCV morphogenesis and maturation. Therefore, MLS000833705 can be used as a molecular probe to study HCV maturation and secretion and possibly guide development of a new class of HCV antivirals.
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Affiliation(s)
- Seung Bum Park
- Liver Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA
| | - Audrey Boyer
- Liver Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA
| | - Zongyi Hu
- Liver Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA
| | - Derek Le
- Liver Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA
| | - T Jake Liang
- Liver Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA
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46
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Sharma G, Raheja H, Das S. Hepatitis C virus: Enslavement of host factors. IUBMB Life 2018; 70:41-49. [PMID: 29281185 DOI: 10.1002/iub.1702] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/06/2017] [Revised: 11/20/2017] [Accepted: 11/28/2017] [Indexed: 12/11/2022]
Abstract
Hepatitis C virus (HCV) has infected over 170 million people world-wide. This infection causes severe liver damage that can progress to hepatocellular carcinoma leading to death of the infected patients. Development of a cell culture model system for the study of HCV infection in the recent past has helped the researchers world-wide to understand the biology of this virus. Studies over the past decade have revealed the tricks played by the virus to sustain itself, for as long as 40 years, in the host setup without being eliminated by the immune system. Today we understand that the host organelles and different cellular proteins are affected during HCV infection. This cytoplasmic virus has all the cellular organelles at its disposal to successfully replicate, from ribosomes and intracellular membranous structures to the nucleus. It modulates these organelles at both the structural and the functional levels. The vast knowledge about the viral genome and viral proteins has also helped in the development of drugs against the virus. Despite the achieved success rate to cure the infected patients, we struggle to eliminate the cases of recurrence and the non-responders. Such cases might emerge owing to the property of the viral genome to accumulate mutations during its succeeding replication cycles which favours its survival. The current situation calls an urgent need for alternate therapeutic strategies to counter this major problem of human health. © 2017 IUBMB Life, 70(1):41-49, 2018.
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Affiliation(s)
- Geetika Sharma
- Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, 560012, Karnataka, India
| | - Harsha Raheja
- Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, 560012, Karnataka, India
| | - Saumitra Das
- Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, 560012, Karnataka, India
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47
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Ashraf MU, Iman K, Khalid MF, Salman HM, Shafi T, Rafi M, Javaid N, Hussain R, Ahmad F, Shahzad-Ul-Hussan S, Mirza S, Shafiq M, Afzal S, Hamera S, Anwar S, Qazi R, Idrees M, Qureshi SA, Chaudhary SU. Evolution of efficacious pangenotypic hepatitis C virus therapies. Med Res Rev 2018; 39:1091-1136. [PMID: 30506705 DOI: 10.1002/med.21554] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/10/2018] [Revised: 10/11/2018] [Accepted: 10/11/2018] [Indexed: 12/12/2022]
Abstract
Hepatitis C compromises the quality of life of more than 350 million individuals worldwide. Over the last decade, therapeutic regimens for treating hepatitis C virus (HCV) infections have undergone rapid advancements. Initially, structure-based drug design was used to develop molecules that inhibit viral enzymes. Subsequently, establishment of cell-based replicon systems enabled investigations into various stages of HCV life cycle including its entry, replication, translation, and assembly, as well as role of host proteins. Collectively, these approaches have facilitated identification of important molecules that are deemed essential for HCV life cycle. The expanded set of putative virus and host-encoded targets has brought us one step closer to developing robust strategies for efficacious, pangenotypic, and well-tolerated medicines against HCV. Herein, we provide an overview of the development of various classes of virus and host-directed therapies that are currently in use along with others that are undergoing clinical evaluation.
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Affiliation(s)
- Muhammad Usman Ashraf
- Biomedical Informatics Research Laboratory, Department of Biology, Lahore University of Management Sciences, Lahore, Pakistan.,Virology Laboratory, Center of Excellence in Molecular Biology, University of the Punjab, Lahore, Pakistan
| | - Kanzal Iman
- Biomedical Informatics Research Laboratory, Department of Biology, Lahore University of Management Sciences, Lahore, Pakistan
| | - Muhammad Farhan Khalid
- Biomedical Informatics Research Laboratory, Department of Biology, Lahore University of Management Sciences, Lahore, Pakistan.,Department of Biomedical Engineering, University of Engineering and Technology, Lahore, Pakistan
| | - Hafiz Muhammad Salman
- Biomedical Informatics Research Laboratory, Department of Biology, Lahore University of Management Sciences, Lahore, Pakistan.,Plant Biotechnology Laboratory, Institute of Agricultural Sciences, University of the Punjab, Lahore, Pakistan
| | - Talha Shafi
- Biomedical Informatics Research Laboratory, Department of Biology, Lahore University of Management Sciences, Lahore, Pakistan
| | - Momal Rafi
- Department of Statistics, University of Gujrat, Gujrat, Pakistan
| | - Nida Javaid
- Department of Biology, Lahore University of Management Sciences, Lahore, Pakistan
| | - Rashid Hussain
- Biomedical Informatics Research Laboratory, Department of Biology, Lahore University of Management Sciences, Lahore, Pakistan
| | - Fayyaz Ahmad
- Department of Statistics, University of Gujrat, Gujrat, Pakistan
| | | | - Shaper Mirza
- Department of Biology, Lahore University of Management Sciences, Lahore, Pakistan
| | - Muhammad Shafiq
- Plant Biotechnology Laboratory, Institute of Agricultural Sciences, University of the Punjab, Lahore, Pakistan
| | - Samia Afzal
- Virology Laboratory, Center of Excellence in Molecular Biology, University of the Punjab, Lahore, Pakistan
| | - Sadia Hamera
- Department of Plant Genetics, Institute of Life Sciences, University of Rostock, Germany
| | - Saima Anwar
- Department of Biomedical Engineering, University of Engineering and Technology, Lahore, Pakistan
| | - Romena Qazi
- Department of Pathology, Shaukat Khanum Memorial Cancer Hospital & Research Centre, Lahore, Pakistan
| | - Muhammad Idrees
- Virology Laboratory, Center of Excellence in Molecular Biology, University of the Punjab, Lahore, Pakistan.,Hazara University, Mansehra, Pakistan
| | - Sohail A Qureshi
- Institute of Integrative Biosciences, CECOS-University of Information Technology and Emerging Sciences, Peshawar, Pakistan
| | - Safee Ullah Chaudhary
- Biomedical Informatics Research Laboratory, Department of Biology, Lahore University of Management Sciences, Lahore, Pakistan
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Boyer A, Park SB, de Boer Y, Li Q, Liang TJ. TM6SF2 Promotes Lipidation and Secretion of Hepatitis C Virus in Infected Hepatocytes. Gastroenterology 2018; 155:1923-1935.e8. [PMID: 30144428 PMCID: PMC6279583 DOI: 10.1053/j.gastro.2018.08.027] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/26/2018] [Revised: 07/17/2018] [Accepted: 08/06/2018] [Indexed: 12/12/2022]
Abstract
BACKGROUND & AIMS Hepatitis C virus (HCV) co-opts the very-low-density lipoprotein pathway for morphogenesis, maturation, and secretion, and circulates as lipoviroparticles (LVPs). We investigated the functions and underlying mechanisms of the lipid-associated TM6SF2 protein in modulating LVP formation and the HCV life cycle. METHODS We knocked down or overexpressed TM6SF2 in hepatic cells and examined HCV infection, measuring viral RNA and protein levels and infectious LVP titers. The density of secreted LVPs was evaluated by iodixanol gradient assay. We measured levels and patterns of TM6SF2 in liver biopsies from 73 patients with chronic hepatitis C, livers of HCV-infected humanized Alb-uPA/SCID/beige mice, and HCV-infected Huh7.5.1 cells. RESULTS TM6SF2 knockdown in hepatocytes reduced viral RNA and infectious viral particle secretion without affecting HCV genome replication, translation, or assembly. Overexpression of TM6SF2 reduced intracellular levels of HCV RNA and infectious LVPs, and conversely increased their levels in the culture supernatants. In HCV-infected cells, TM6SF2 overexpression resulted in production of more infectious LVPs in the lower-density fractions of supernatant. HCV infection increased TM6SF2 expression in cultured cells, humanized livers of mice, and liver tissues of HCV patients. TM6SF2 messenger RNA levels correlated positively with HCV RNA levels in liver biopsies from patients. SREBF2 appears to mediate the ability of HCV to increase the expression of TM6SF2 in hepatic cells. CONCLUSIONS In studies of cells, mice and human liver tissues, we found TM6SF2 is required for maturation, lipidation, and secretion of infectious LVPs. HCV, in turn, up-regulates expression of TM6SF2 to facilitate productive infection.
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Murayama A, Saitoh H, Takeuchi A, Yamada N, Matsumura T, Shiina M, Muramatsu M, Wakita T, Imawari M, Kato T. Vitamin D derivatives inhibit hepatitis C virus production through the suppression of apolipoprotein. Antiviral Res 2018; 160:55-63. [PMID: 30339849 DOI: 10.1016/j.antiviral.2018.10.014] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/01/2018] [Revised: 09/21/2018] [Accepted: 10/14/2018] [Indexed: 01/28/2023]
Abstract
Supplementation with vitamin D (VD) has been reported to improve the efficacy of interferon-based therapy for chronic hepatitis C. We found that 25-hydroxyvitamin D3 (25-(OH)D3), one of the metabolites of VD, has antiviral effects by inhibiting the infectious virus production of the hepatitis C virus (HCV). In this study, to clarify the underlying mechanisms of the anti-HCV effects, we searched VD derivatives that have anti-HCV effects and identified the common target molecule in the HCV life cycle by using an HCV cell culture system. After infection of Huh-7.5.1 cells with cell culture-generated HCV, VD derivatives were added to culture media, and the propagation of HCV was assessed by measuring the HCV core antigen levels in culture media and cell lysates. To determine the step in the HCV life cycle affected by these compounds, the single-cycle virus production assay was used with a CD81-negative cell line. Of the 14 structural derivatives of VD, an anti-HCV effect was detected in 9 compounds. Cell viability was not affected by these effective compounds. The 2 representative VD derivatives inhibited the infectious virus production in the single-cycle virus production assay. Treatment with these compounds and 25-(OH)D3 suppressed the expression of apolipoprotein A1 and C3, which are known to be involved in infectious virus production of HCV, and the knockdown of these apolipoproteins reduced infectious virus production. In conclusion, we identified several compounds with anti-HCV activity by screening VD derivatives. These compounds reduce the infectious virus production of HCV by suppressing the expression of apolipoproteins in host cells.
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Affiliation(s)
- Asako Murayama
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Hiroshi Saitoh
- Teijin Institute for Bio-medical Research, Teijin Pharma Ltd., Tokyo, Japan
| | - Akiko Takeuchi
- Teijin Institute for Bio-medical Research, Teijin Pharma Ltd., Tokyo, Japan
| | - Norie Yamada
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Takuya Matsumura
- Division of Gastroenterology, Department of Medicine, Showa University School of Medicine, Tokyo, Japan
| | - Masaaki Shiina
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan; Department of Gastroenterology and Hepatology, Shin-Yurigaoka General Hospital, Kawasaki, Japan
| | - Masamichi Muramatsu
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Takaji Wakita
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan
| | - Michio Imawari
- Research Institute for Gastrointestinal and Liver Diseases, Shin-Yurigaoka General Hospital, Kawasaki, Japan
| | - Takanobu Kato
- Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan.
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50
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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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