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Izumida K, Hara Y, Iwatsuki R, Ohta S, Tabata K, Morita E. In vitro characteristics of purified recombinant hepatitis C virus core protein. Virology 2025; 601:110297. [PMID: 39536644 DOI: 10.1016/j.virol.2024.110297] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/16/2024] [Revised: 10/18/2024] [Accepted: 11/07/2024] [Indexed: 11/16/2024]
Abstract
In our previous study, we established a method for purifying bacterially expressed HCV core 1-164 under non-denaturing conditions. In the present study, we elucidated the characteristics of the purified core. The purified HCV core exhibited a notable affinity for HCV RNA, with a Kd of 3 nM, as determined by a filter binding assay. Electron microscopy analysis revealed that the purified HCV core self-assembled with RNA into spherical structures approximately 50 nm in diameter. Additionally, we demonstrated the direct binding of the purified HCV core to the purified endoplasmic reticulum (ER). Moreover, lipid strip assays revealed specific binding of the purified HCV core to specific phospholipids, suggesting that the core plays a role in specific ER membrane domains. These studies reveal the biochemical characteristics of the HCV core that significantly advance our understanding of its role as a capsid protein in the viral lifecycle and pathogenesis.
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Affiliation(s)
- Kyo Izumida
- Laboratory of Viral Infection, Research Institute for Microbial Diseases, Osaka University, Suita, 565-0871, Japan
| | - Yumiko Hara
- Laboratory of Viral Infection, Research Institute for Microbial Diseases, Osaka University, Suita, 565-0871, Japan; Department of Biochemistry and Molecular Biology, Faculty of Agriculture and Life Science, Hirosaki University, Hirosaki, 036-8561, Japan
| | - Ryuta Iwatsuki
- Department of Biochemistry and Molecular Biology, Faculty of Agriculture and Life Science, Hirosaki University, Hirosaki, 036-8561, Japan
| | - Sora Ohta
- Department of Biochemistry and Molecular Biology, Faculty of Agriculture and Life Science, Hirosaki University, Hirosaki, 036-8561, Japan
| | - Keisuke Tabata
- Laboratory of Viral Infection, Research Institute for Microbial Diseases, Osaka University, Suita, 565-0871, Japan; Laboratory of Intracellular Membrane Dynamics, Graduate School of Frontier Biosciences Osaka University, Suita, 565-0871, Japan; Department of Genetics, Graduate School of Medicine Osaka University, Suita, 565-0871, Japan
| | - Eiji Morita
- Laboratory of Viral Infection, Research Institute for Microbial Diseases, Osaka University, Suita, 565-0871, Japan; Department of Biochemistry and Molecular Biology, Faculty of Agriculture and Life Science, Hirosaki University, Hirosaki, 036-8561, Japan.
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Nepal S, Holmstrom ED. Single-molecule-binding studies of antivirals targeting the hepatitis C virus core protein. J Virol 2023; 97:e0089223. [PMID: 37772835 PMCID: PMC10617558 DOI: 10.1128/jvi.00892-23] [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: 06/14/2023] [Accepted: 08/10/2023] [Indexed: 09/30/2023] Open
Abstract
IMPORTANCE The hepatitis C virus is associated with nearly 300,000 deaths annually. At the core of the virus is an RNA-protein complex called the nucleocapsid, which consists of the viral genome and many copies of the core protein. Because the assembly of the nucleocapsid is a critical step in viral replication, a considerable amount of effort has been devoted to identifying antiviral therapeutics that can bind to the core protein and disrupt assembly. Although several candidates have been identified, little is known about how they interact with the core protein or how those interactions alter the structure and thus the function of this viral protein. Our work biochemically characterizes several of these binding interactions, highlighting both similarities and differences as well as strengths and weaknesses. These insights bolster the notion that this viral protein is a viable target for novel therapeutics and will help to guide future developments of these candidate antivirals.
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Affiliation(s)
- Sudip Nepal
- Department of Molecular Biosciences, University of Kansas, Lawrence, Kansas, USA
| | - Erik D. Holmstrom
- Department of Molecular Biosciences, University of Kansas, Lawrence, Kansas, USA
- Department of Chemistry, University of Kansas, Lawrence, Kansas, USA
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Duponchel S, Monnier L, Molle J, Bendridi N, Alam MR, Gaballah A, Grigorov B, Ivanov A, Schmiel M, Odenthal M, Ovize M, Rieusset J, Zoulim F, Bartosch B. Hepatitis C virus replication requires integrity of mitochondria-associated ER membranes. JHEP REPORTS : INNOVATION IN HEPATOLOGY 2022; 5:100647. [PMID: 36718430 PMCID: PMC9883273 DOI: 10.1016/j.jhepr.2022.100647] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 10/13/2022] [Revised: 11/15/2022] [Accepted: 11/21/2022] [Indexed: 12/13/2022]
Abstract
Background & Aims Chronic HCV infection causes cellular stress, fibrosis and predisposes to hepatocarcinogenesis. Mitochondria play key roles in orchestrating stress responses by regulating bioenergetics, inflammation and apoptosis. To better understand the role of mitochondria in the viral life cycle and disease progression of chronic hepatitis C, we studied morphological and functional mitochondrial alterations induced by HCV using productively infected hepatoma cells and patient livers. Methods Biochemical and imaging assays were used to assess localization of cellular and viral proteins and mitochondrial functions in cell cultures and liver biopsies. Cyclophilin D (CypD) knockout was performed using CRISPR/Cas9 technology. Viral replication was quantified by quantitative reverse-transcription PCR and western blotting. Results Several HCV proteins were found to associate with mitochondria-associated endoplasmic reticulum (ER) membranes (MAMs), the points of contact between the ER and mitochondria. Downregulation of CypD, which is known to disrupt MAM integrity, reduced viral replication, suggesting that MAMs play an important role in the viral life cycle. This process was rescued by ectopic CypD expression. Furthermore, HCV proteins were found to associate with voltage dependent anion channel 1 (VDAC1) at MAMs and to reduce VDAC1 protein levels at MAMs in vitro and in patient biopsies. This association did not affect MAM-associated functions in glucose homeostasis and Ca2+ signaling. Conclusions HCV proteins associate specifically with MAMs and MAMs play an important role in viral replication. The association between viral proteins and MAMs did not impact Ca2+ signaling between the ER and mitochondria or glucose homeostasis. Whether additional functions of MAMs and/or VDAC are impacted by HCV and contribute to the associated pathology remains to be assessed. Impact and implications Hepatitis C virus infects the liver, where it causes inflammation, cell damage and increases the long-term risk of liver cancer. We show that several HCV proteins interact with mitochondria in liver cells and alter the composition of mitochondrial subdomains. Importantly, HCV requires the architecture of these mitochondrial subdomains to remain intact for efficient viral replication.
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Key Words
- CypD, cyclophilin D
- DMVs, double membrane vesicles
- EM, electron microscopy
- ER, endoplasmic reticulum
- Grp75, glucose-regulated protein 75
- HCC, hepatocellular carcinoma
- HCVcc, cell culture-derived HCV
- IP, immunoprecipitation
- IP3R1, inositol trisphosphate receptor 1
- KO, knockout
- MAMs, mitochondria-associated ER membranes
- MOI, multiplicity of infection
- OMM, outer mitochondrial membrane
- PLA, proximity ligation assay
- S1R, sigma 1 receptor
- VDAC, voltage-dependent anion channel
- dpi, days post infection
- fibrosis
- hepatitis C virus
- mitochondria-associated ER membranes
- voltage-dependent anion channel 1
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Affiliation(s)
- Sarah Duponchel
- Univ Lyon, Université Claude Bernard Lyon 1, INSERM 1052, CNRS 5286, Centre Léon Bérard, Centre de Recherche en Cancérologie de Lyon, Lyon, 69434, France
| | - Lea Monnier
- Univ Lyon, Université Claude Bernard Lyon 1, INSERM 1052, CNRS 5286, Centre Léon Bérard, Centre de Recherche en Cancérologie de Lyon, Lyon, 69434, France
| | - Jennifer Molle
- Univ Lyon, Université Claude Bernard Lyon 1, INSERM 1052, CNRS 5286, Centre Léon Bérard, Centre de Recherche en Cancérologie de Lyon, Lyon, 69434, France
| | - Nadia Bendridi
- Laboratoire CarMeN, INSERM U-1060, INRA U-1397, Université Lyon, Université Claude Bernard Lyon 1, Pierre Bénite, 69495, France
| | - Muhammad Rizwan Alam
- CarMeN Laboratory, Hôpital Louis Pradel, Hospices Civils de Lyon, Université de Lyon and Explorations Fonctionnelles Cardiovasculaires, INSERM U1060, Lyon, France
| | - Ahmed Gaballah
- Univ Lyon, Université Claude Bernard Lyon 1, INSERM 1052, CNRS 5286, Centre Léon Bérard, Centre de Recherche en Cancérologie de Lyon, Lyon, 69434, France,Microbiology Department, Medical Research Institute, Alexandria University, Egypt
| | - Boyan Grigorov
- Univ Lyon, Université Claude Bernard Lyon 1, INSERM 1052, CNRS 5286, Centre Léon Bérard, Centre de Recherche en Cancérologie de Lyon, Lyon, 69434, France
| | - Alexander Ivanov
- Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russia
| | - Marcel Schmiel
- Institute of Pathology, University Hospital of Cologne and Center for Molecular Medicine (CMMC), University of Cologne, Germany
| | - Margarete Odenthal
- Institute of Pathology, University Hospital of Cologne and Center for Molecular Medicine (CMMC), University of Cologne, Germany
| | - Michel Ovize
- CarMeN Laboratory, Hôpital Louis Pradel, Hospices Civils de Lyon, Université de Lyon and Explorations Fonctionnelles Cardiovasculaires, INSERM U1060, Lyon, France
| | - Jennifer Rieusset
- Laboratoire CarMeN, INSERM U-1060, INRA U-1397, Université Lyon, Université Claude Bernard Lyon 1, Pierre Bénite, 69495, France
| | - Fabien Zoulim
- Univ Lyon, Université Claude Bernard Lyon 1, INSERM 1052, CNRS 5286, Centre Léon Bérard, Centre de Recherche en Cancérologie de Lyon, Lyon, 69434, France,Hospices Civils de Lyon, France
| | - Birke Bartosch
- Univ Lyon, Université Claude Bernard Lyon 1, INSERM 1052, CNRS 5286, Centre Léon Bérard, Centre de Recherche en Cancérologie de Lyon, Lyon, 69434, France,Corresponding author. Address: Cancer Research Center Lyon, 151 cours Albert Thomas, 69434 Lyon, France; Tel.: 0033472681975, fax: 0033472681971
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Viral Infection Modulates Mitochondrial Function. Int J Mol Sci 2021; 22:ijms22084260. [PMID: 33923929 PMCID: PMC8073244 DOI: 10.3390/ijms22084260] [Citation(s) in RCA: 21] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/30/2021] [Revised: 04/14/2021] [Accepted: 04/15/2021] [Indexed: 02/08/2023] Open
Abstract
Mitochondria are important organelles involved in metabolism and programmed cell death in eukaryotic cells. In addition, mitochondria are also closely related to the innate immunity of host cells against viruses. The abnormality of mitochondrial morphology and function might lead to a variety of diseases. A large number of studies have found that a variety of viral infections could change mitochondrial dynamics, mediate mitochondria-induced cell death, and alter the mitochondrial metabolic status and cellular innate immune response to maintain intracellular survival. Meanwhile, mitochondria can also play an antiviral role during viral infection, thereby protecting the host. Therefore, mitochondria play an important role in the interaction between the host and the virus. Herein, we summarize how viral infections affect microbial pathogenesis by altering mitochondrial morphology and function and how viruses escape the host immune response.
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Bai Z, Zhao X, Li C, Sheng C, Li H. EV71 virus reduces Nrf2 activation to promote production of reactive oxygen species in infected cells. Gut Pathog 2020; 12:22. [PMID: 32346399 PMCID: PMC7181592 DOI: 10.1186/s13099-020-00361-w] [Citation(s) in RCA: 24] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/24/2019] [Accepted: 04/15/2020] [Indexed: 12/13/2022] Open
Abstract
BACKGROUND Emerging evidence closely links Enterovirus 71 (EV71) infection with the generation of reactive oxygen species (ROS). Excess ROS results in apoptosis and exacerbates inflammatory reactions. The Keap1-Nrf2 axis serves as an essential oxidant counteracting pathway. METHODS The present study aimed to elucidate the role of the Keap1-Nrf2 pathway in modulating apoptosis and inflammatory reactions triggered by oxidative stress in Vero and RD cells upon EV71 infection. RESULTS Elevated ROS production was identified in EV71 infected Vero and RD cells. The percentage of dead cells and expression of inflammation-promoting cytokines were increased in these cells. EV71 infected cells also displayed reinforced Keap1 expression and abrogated Nrf2 expression. Keap1 silencing resulted in the downstream aggregation of the Nrf2 protein and heme oxygenase-1 HO-1. Keap1 silencing repressed ubiquitination and reinforced Nrf2 nuclear trafficking. Furthermore, silencing Keap1 expression repressed ROS production, cell death, and inflammatory reactions in EV71 infected RD and Vero cells. In contrast, silencing of both Keap1 and Nrf2 restored ROS production, cell death, and inflammatory reactions. Nrf2 and Keap1 modulated the stimulation of the Akt sensor and extrinsic as well as intrinsic cell death pathways, resulting in EV71-triggered cell death and inflammatory reactions. CONCLUSIONS EV71 infection can trigger ROS production, cell death, and inflammatory reactions by modulating the Nrf2 and Keap1 levels of infected cells.
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Affiliation(s)
- Zhenzi Bai
- Infectious Department, China-Japan Union Hospital, Jilin University, No.126, Xiantai Street, Economic Development Zone, Changchun, 130033 Jilin China
| | - Xiaonan Zhao
- Infectious Department, China-Japan Union Hospital, Jilin University, No.126, Xiantai Street, Economic Development Zone, Changchun, 130033 Jilin China
| | - Chenghua Li
- Infectious Department, China-Japan Union Hospital, Jilin University, No.126, Xiantai Street, Economic Development Zone, Changchun, 130033 Jilin China
| | - Chuanlun Sheng
- Infectious Department, China-Japan Union Hospital, Jilin University, No.126, Xiantai Street, Economic Development Zone, Changchun, 130033 Jilin China
| | - Hongyan Li
- Infectious Department, China-Japan Union Hospital, Jilin University, No.126, Xiantai Street, Economic Development Zone, Changchun, 130033 Jilin China
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Li H, Bai Z, Li C, Sheng C, Zhao X. EV71 infection induces cell apoptosis through ROS generation and SIRT1 activation. J Cell Biochem 2020; 121:4321-4331. [PMID: 31898369 DOI: 10.1002/jcb.29628] [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] [Received: 07/31/2019] [Accepted: 12/19/2019] [Indexed: 12/28/2022]
Abstract
Several studies have substantiated the correlation between reactive oxygen species (ROS) and Sirtuin 1 (SIRT1). Normally, enterovirus 71 (EV71) is associated with severe clinical manifestations and death. However, the effect of EV71 on the induction of cellular death and the interplay between ROS/SIRT1 in cell death has not been confirmed yet. In the current study, an increase in the number of apoptotic cells was observed as soon as the EV71 infection was initiated in cells and mice. Furthermore, EV71 infection also promoted a rise in the levels of three commonly known proinflammatory cytokines, interleukin 1β (IL-1β), IL-6, and tumor necrosis factor-α. During EV71-induced apoptosis in the different cell lines, ROS generation and SIRT1 downregulation were observed. Further investigations showed that the administration of ROS inhibitor, N-acetyl- l-cysteine (NAC), reduced the level of apoptosis and inflammation, reduced EV71 propagation, and increased SIRT1 expression in EV71-infected cells. In addition, combined administration of NAC and EX527 (SIRT1 inhibitor) restored apoptosis in the EV71-infected cells, which was reduced due to NAC. This data demonstrated that ROS generation is positively associated with EV71-induced apoptosis and inflammation, while this effect could be reversed by SIRT1 inhibition. Collectively, we have shown that EV71 induces apoptosis and inflammation by promoting ROS generation and reducing SIRT1 expression.
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Affiliation(s)
- Hongyan Li
- Infectious Department, China-Japan Union Hospital, Jilin University, Changchun, China
| | - Zhenzi Bai
- Infectious Department, China-Japan Union Hospital, Jilin University, Changchun, China
| | - Chenghua Li
- Infectious Department, China-Japan Union Hospital, Jilin University, Changchun, China
| | - Chuanlun Sheng
- Infectious Department, China-Japan Union Hospital, Jilin University, Changchun, China
| | - Xiaonan Zhao
- Infectious Department, China-Japan Union Hospital, Jilin University, Changchun, China
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Bender D, Hildt E. Effect of Hepatitis Viruses on the Nrf2/Keap1-Signaling Pathway and Its Impact on Viral Replication and Pathogenesis. Int J Mol Sci 2019; 20:ijms20184659. [PMID: 31546975 PMCID: PMC6769940 DOI: 10.3390/ijms20184659] [Citation(s) in RCA: 29] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/29/2019] [Revised: 09/16/2019] [Accepted: 09/17/2019] [Indexed: 12/15/2022] Open
Abstract
With respect to their genome and their structure, the human hepatitis B virus (HBV) and hepatitis C virus (HCV) are complete different viruses. However, both viruses can cause an acute and chronic infection of the liver that is associated with liver inflammation (hepatitis). For both viruses chronic infection can lead to fibrosis, cirrhosis and hepatocellular carcinoma (HCC). Reactive oxygen species (ROS) play a central role in a variety of chronic inflammatory diseases. In light of this, this review summarizes the impact of both viruses on ROS-generating and ROS-inactivating mechanisms. The focus is on the effect of both viruses on the transcription factor Nrf2 (nuclear factor erythroid 2 (NF-E2)-related factor 2). By binding to its target sequence, the antioxidant response element (ARE), Nrf2 triggers the expression of a variety of cytoprotective genes including ROS-detoxifying enzymes. The review summarizes the literature about the pathways for the modulation of Nrf2 that are deregulated by HBV and HCV and describes the impact of Nrf2 deregulation on the viral life cycle of the respective viruses and the virus-associated pathogenesis.
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Affiliation(s)
- Daniela Bender
- Department of Virology, Paul-Ehrlich-Institut, Paul-Ehrlich-Straβe 51-59, D-63225 Langen, Germany.
| | - Eberhard Hildt
- Department of Virology, Paul-Ehrlich-Institut, Paul-Ehrlich-Straβe 51-59, D-63225 Langen, Germany.
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Naringenin ameliorates insulin resistance by modulating endoplasmic reticulum stress in hepatitis C virus-infected liver. Biomed Pharmacother 2019; 115:108848. [PMID: 31039496 DOI: 10.1016/j.biopha.2019.108848] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/01/2019] [Revised: 04/02/2019] [Accepted: 04/02/2019] [Indexed: 12/15/2022] Open
Abstract
Hepatitis C virus (HCV) infection may lead to hepatic insulin resistance (IR), and endoplasmic reticulum (ER) stress has been found to induce IR. In our previous study, naringenin (NGEN) had an insulin sensitization effect on the HCV core protein (HCVCP) infected mouse livers. In the present study, we examined the effects of NGEN on HCVCP infection-induced ER stress and investigated the insulin sensitization mechanism involved. We found that XBP1s was up-regulated in the livers of HCV-infected patients, in hepatocytes with HCV infection, and in HCVCP-infected mice. HCVCP induces ER stress in the mouse liver and hepatocytes by increasing XBP1s and downstream gene expression. Pre-treatment with NGEN inhibited the ER stress and downstream gene expression both in vivo and in vitro. Similar to the HCVCP infection results, NGEN also inhibited the ER stress in tunicamycin-treated Huh-7.5.1 cells. In addition, the role of IRE1α in HCVCP-induced IR was detected, and knockdown of IRE1α abolished HCVCP-stimulated IR. Overexpression induced IR but could be abolished by NGEN. NGEN also blocked the HCVCP-induced IRE1α expression levels that were up-regulated in vivo. Our data reveal that ER stress may be associated with HCV-induced IR, and NGEN treatment inhibited ER stress activity and increased insulin sensitivity by decreasing the expression of IRE1α.
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USP15 Participates in Hepatitis C Virus Propagation through Regulation of Viral RNA Translation and Lipid Droplet Formation. J Virol 2019; 93:JVI.01708-18. [PMID: 30626683 DOI: 10.1128/jvi.01708-18] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/30/2018] [Accepted: 12/23/2018] [Indexed: 12/12/2022] Open
Abstract
Hepatitis C virus (HCV) utilizes cellular factors for efficient propagation. Ubiquitin is covalently conjugated to the substrate to alter its stability or to modulate signal transduction. In this study, we examined the importance of ubiquitination for HCV propagation. We found that inhibition of deubiquitinating enzymes (DUBs) or overexpression of nonspecific DUBs impaired HCV replication, suggesting that ubiquitination regulates HCV replication. To identify specific DUBs involved in HCV propagation, we set up RNA interference (RNAi) screening against DUBs and successfully identified ubiquitin-specific protease 15 (USP15) as a novel host factor for HCV propagation. Our studies showed that USP15 is involved in translation of HCV RNA and production of infectious HCV particles. In addition, deficiency of USP15 in human hepatic cell lines (Huh7 and Hep3B/miR-122 cells) but not in a nonhepatic cell line (293T cells) impaired HCV propagation, suggesting that USP15 participates in HCV propagation through the regulation of hepatocyte-specific functions. Moreover, we showed that loss of USP15 had no effect on innate immune responses in vitro and in vivo We also found that USP15-deficient Huh7 cells showed reductions in the amounts of lipid droplets (LDs), and the addition of palmitic acids restored the production of infectious HCV particles. Taken together, these data suggest that USP15 participates in HCV propagation by regulating the translation of HCV RNA and the formation of LDs.IMPORTANCE Although ubiquitination has been shown to play important roles in the HCV life cycle, the roles of deubiquitinating enzymes (DUBs), which cleave ubiquitin chains from their substrates, in HCV propagation have not been investigated. Here, we identified USP15 as a DUB regulating HCV propagation. USP15 showed no interaction with viral proteins and no participation in innate immune responses. Deficiency of USP15 in Huh7 cells resulted in suppression of the translation of HCV RNA and reduction in the amounts of lipid droplets, and the addition of fatty acids partially restored the production of infectious HCV particles. These data suggest that USP15 participates in HCV propagation in hepatic cells through the regulation of viral RNA translation and lipid metabolism.
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Hino K, Nishina S, Sasaki K, Hara Y. Mitochondrial damage and iron metabolic dysregulation in hepatitis C virus infection. Free Radic Biol Med 2019; 133:193-199. [PMID: 30268888 DOI: 10.1016/j.freeradbiomed.2018.09.044] [Citation(s) in RCA: 30] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/15/2018] [Revised: 09/25/2018] [Accepted: 09/26/2018] [Indexed: 02/06/2023]
Abstract
Hepatitis C virus (HCV) infection often leads to chronic hepatitis that can progress to liver cirrhosis and hepatocellular carcinoma (HCC). Although HCV infection is expected to decrease due to the high rate of HCV eradication via the rapid dissemination and use of directly acting antivirals, HCV infection remains a leading cause of HCC. Although the mechanisms underlying the HCC development are not fully understood, oxidative stress is present to a greater degree in HCV infection than in other inflammatory liver diseases and has been proposed as a major mechanism of liver injury in patients with chronic hepatitis C. Hepatocellular mitochondrial alterations and iron accumulation are well-known characteristics in patients with chronic hepatitis C and are closely related to oxidative stress, since the mitochondria are the main site of reactive oxygen species generation, and iron produces hydroxy radicals via the Fenton reaction. In addition, phlebotomy is an iron reduction approach that aims to lower serum transaminase levels in patients with chronic hepatitis C. Here, we review and discuss the mechanisms by which HCV induces mitochondrial damage and iron accumulation in the liver and offer new insights concerning how mitochondrial damage and iron accumulation are linked to the development of HCC.
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Affiliation(s)
- Keisuke Hino
- Department of Hepatology and Pancreatology, Kawasaki Medical School, 577 Matsushima Kurashiki, Okayama 701-0192, Japan.
| | - Sohij Nishina
- Department of Hepatology and Pancreatology, Kawasaki Medical School, 577 Matsushima Kurashiki, Okayama 701-0192, Japan.
| | - Kyo Sasaki
- Department of Hepatology and Pancreatology, Kawasaki Medical School, 577 Matsushima Kurashiki, Okayama 701-0192, Japan.
| | - Yuichi Hara
- Department of Hepatology and Pancreatology, Kawasaki Medical School, 577 Matsushima Kurashiki, Okayama 701-0192, Japan.
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Jansons J, Sominskaya I, Petrakova N, Starodubova ES, Smirnova OA, Alekseeva E, Bruvere R, Eliseeva O, Skrastina D, Kashuba E, Mihailova M, Kochetkov SN, Ivanov AV, Isaguliants MG. The Immunogenicity in Mice of HCV Core Delivered as DNA Is Modulated by Its Capacity to Induce Oxidative Stress and Oxidative Stress Response. Cells 2019; 8:cells8030208. [PMID: 30823485 PMCID: PMC6468923 DOI: 10.3390/cells8030208] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/22/2018] [Revised: 02/06/2019] [Accepted: 02/20/2019] [Indexed: 12/16/2022] Open
Abstract
HCV core is an attractive HCV vaccine target, however, clinical or preclinical trials of core-based vaccines showed little success. We aimed to delineate what restricts its immunogenicity and improve immunogenic performance in mice. We designed plasmids encoding full-length HCV 1b core and its variants truncated after amino acids (aa) 60, 98, 152, 173, or up to aa 36 using virus-derived or synthetic polynucleotides (core191/60/98/152/173/36_191v or core152s DNA, respectively). We assessed their level of expression, route of degradation, ability to trigger the production of reactive oxygen species/ROS, and to activate the components of the Nrf2/ARE antioxidant defense pathway heme oxygenase 1/HO-1 and NAD(P)H: quinone oxidoreductase/Nqo-1. All core variants with the intact N-terminus induced production of ROS, and up-regulated expression of HO-1 and Nqo-1. The capacity of core variants to induce ROS and up-regulate HO-1 and Nqo-1 expression predetermined their immunogenicity in DNA-immunized BALB/c and C57BL/6 mice. The most immunogenic was core 152s, expressed at a modest level and inducing moderate oxidative stress and oxidative stress response. Thus, immunogenicity of HCV core is shaped by its ability to induce ROS and oxidative stress response. These considerations are important in understanding the mechanisms of viral suppression of cellular immune response and in HCV vaccine design.
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Affiliation(s)
- Juris Jansons
- Department of Pathology, Riga Stradins University, LV-1007 Riga, Latvia.
- Latvian Biomedical Research and Study Centre, LV-1067 Riga, Latvia.
- Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, SE-171 77 Stockholm, Sweden.
| | - Irina Sominskaya
- Latvian Biomedical Research and Study Centre, LV-1067 Riga, Latvia.
| | - Natalia Petrakova
- N.F. Gamaleya Research Center of Epidemiology and Microbiology, Ministry of Health of the Russian Federation, 123098 Moscow, Russia.
| | - Elizaveta S Starodubova
- N.F. Gamaleya Research Center of Epidemiology and Microbiology, Ministry of Health of the Russian Federation, 123098 Moscow, Russia.
- Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, 119991 Moscow, Russia.
| | - Olga A Smirnova
- Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, 119991 Moscow, Russia.
| | - Ekaterina Alekseeva
- Department of Pathology, Riga Stradins University, LV-1007 Riga, Latvia.
- Latvian Biomedical Research and Study Centre, LV-1067 Riga, Latvia.
| | - Ruta Bruvere
- Latvian Biomedical Research and Study Centre, LV-1067 Riga, Latvia.
| | - Olesja Eliseeva
- N.F. Gamaleya Research Center of Epidemiology and Microbiology, Ministry of Health of the Russian Federation, 123098 Moscow, Russia.
| | - Dace Skrastina
- Department of Pathology, Riga Stradins University, LV-1007 Riga, Latvia.
- Latvian Biomedical Research and Study Centre, LV-1067 Riga, Latvia.
| | - Elena Kashuba
- Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, SE-171 77 Stockholm, Sweden.
- RE Kavetsky Institite of Experimental Pathology, Oncology and Radiobiology, The National Academy of Sciences of Ukraine, 03022 Kyiv, Ukraine.
| | - Marija Mihailova
- Latvian Biomedical Research and Study Centre, LV-1067 Riga, Latvia.
| | - Sergey N Kochetkov
- Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, 119991 Moscow, Russia.
| | - Alexander V Ivanov
- Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, 119991 Moscow, Russia.
| | - Maria G Isaguliants
- Department of Pathology, Riga Stradins University, LV-1007 Riga, Latvia.
- Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, SE-171 77 Stockholm, Sweden.
- N.F. Gamaleya Research Center of Epidemiology and Microbiology, Ministry of Health of the Russian Federation, 123098 Moscow, Russia.
- MP Chumakov Center for Research and Development of Immune and Biological Preparations of RAS, 108819 Moscow, Russia.
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12
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Zhang C, Liu P. The New Face of the Lipid Droplet: Lipid Droplet Proteins. Proteomics 2018; 19:e1700223. [DOI: 10.1002/pmic.201700223] [Citation(s) in RCA: 49] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/31/2018] [Revised: 08/13/2018] [Indexed: 12/20/2022]
Affiliation(s)
- Congyan Zhang
- National Laboratory of BiomacromoleculesCAS Center for Excellence in BiomacromoleculesInstitute of BiophysicsChinese Academy of Sciences Beijing 100101 China
- University of Chinese Academy of Sciences Beijing 100049 China
| | - Pingsheng Liu
- National Laboratory of BiomacromoleculesCAS Center for Excellence in BiomacromoleculesInstitute of BiophysicsChinese Academy of Sciences Beijing 100101 China
- University of Chinese Academy of Sciences Beijing 100049 China
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13
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Mansouri A, Gattolliat CH, Asselah T. Mitochondrial Dysfunction and Signaling in Chronic Liver Diseases. Gastroenterology 2018; 155:629-647. [PMID: 30012333 DOI: 10.1053/j.gastro.2018.06.083] [Citation(s) in RCA: 515] [Impact Index Per Article: 73.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/05/2017] [Revised: 05/23/2018] [Accepted: 06/10/2018] [Indexed: 12/12/2022]
Abstract
Mitochondria regulate hepatic lipid metabolism and oxidative stress. Ultrastructural mitochondrial lesions, altered mitochondrial dynamics, decreased activity of respiratory chain complexes, and impaired ability to synthesize adenosine triphosphate are observed in liver tissues from patients with alcohol-associated and non-associated liver diseases. Increased lipogenesis with decreased fatty acid β-oxidation leads to the accumulation of triglycerides in hepatocytes, which, combined with increased levels of reactive oxygen species, contributes to insulin resistance in patients with steatohepatitis. Moreover, mitochondrial reactive oxygen species mediate metabolic pathway signaling; alterations in these pathways affect development and progression of chronic liver diseases. Mitochondrial stress and lesions promote cell death, liver fibrogenesis, inflammation, and the innate immune responses to viral infections. We review the involvement of mitochondrial processes in development of chronic liver diseases, such as nonalcoholic fatty, alcohol-associated, and drug-associated liver diseases, as well as hepatitis B and C, and discuss how they might be targeted therapeutically.
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Affiliation(s)
- Abdellah Mansouri
- Centre de Recherche sur l'Inflammation, Institut National de la Santé et de la Recherche Médicale, Unité Mixte de Recherche 1149, Université Paris Diderot, PRES Paris Sorbonne Cité, Paris, France
| | - Charles-Henry Gattolliat
- Centre de Recherche sur l'Inflammation, Institut National de la Santé et de la Recherche Médicale, Unité Mixte de Recherche 1149, Université Paris Diderot, PRES Paris Sorbonne Cité, Paris, France
| | - Tarik Asselah
- Centre de Recherche sur l'Inflammation, Institut National de la Santé et de la Recherche Médicale, Unité Mixte de Recherche 1149, Université Paris Diderot, PRES Paris Sorbonne Cité, Paris, France; Department of Hepatology, Assistance Publique-Hôpitaux de Paris, Hôpital Beaujon, Clichy, France.
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14
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Jia B, Yu D, Yu G, Cheng Y, Wang Y, Yi X, Li X, Wang Y. Naringenin improve hepatitis C virus infection induced insulin resistance by increase PTEN expression via p53-dependent manner. Biomed Pharmacother 2018; 103:746-754. [PMID: 29684853 DOI: 10.1016/j.biopha.2018.04.110] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/23/2018] [Revised: 04/13/2018] [Accepted: 04/13/2018] [Indexed: 02/07/2023] Open
Abstract
Hepatitis C virus (HCV) infection may finally lead to hepatocellular carcinoma (HCC), and also associated with insulin resistance (IR). Naringenin (NGEN), a flavonoid found in grapefruit, has anti-virus, anti-inflammation and insulin sensitization effects. In the present study we examined the effects of NGEN on HCV core protein (HCVCP) infection induced IR and investigated the mechanism involved. We found that NGEN ameliorated IR and glucose tolerance in HCVCP infected mice by increase the phosphorylation of Akt at both Ser473 and Thr308 site, and also inhibited the inflammation cytokine production and T-cell immune response. Similar to the in vivo results, NGEN also improved the insulin response and showed anti-inflammation effect in HCVCP infected Huh-7.5.1 cells. In addition, NGEN up-regulated the phosphatase and tensin homolog deleted on chromosome ten (PTEN) both in protein and mRNA levels. Furthermore, overexpress of PTEN abolished the HCVCP-stimulated IR and decreased the inflammation cytokine release. NGEN also blocked the interaction between HCVCP and p53, upregulated the endogenous p21/waf1 expression which indiacting the activation of p53. The p53 wild type could upregulate NGEN-stimulated PTEN expression while R273H mut-p53 showed no similar function. Our data reveals that NGEN increases insulin sensitivity in HCVCP infected liver by up-regulating PTEN in p53-dependent manner.
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Affiliation(s)
- Benli Jia
- Department of General Surgery, The Second Affiliated Hospital of Anhui Medical University, Hefei, Anhui, 230601, China
| | - Dongsheng Yu
- Department of Chinese Medicine, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, 450052, China; Department of Pharmacology, School of Basic Medical Science, Nanjing Medical University, 101 Longmian Rd, Nanjing, Jiangsu, 211166, China
| | - Gang Yu
- Department of General Surgery, The Second Affiliated Hospital of Anhui Medical University, Hefei, Anhui, 230601, China
| | - Yunsheng Cheng
- Department of General Surgery, The Second Affiliated Hospital of Anhui Medical University, Hefei, Anhui, 230601, China
| | - Yang Wang
- Department of General Surgery, The Second Affiliated Hospital of Anhui Medical University, Hefei, Anhui, 230601, China
| | - Xiaoqiang Yi
- Department of General Surgery, The Second Affiliated Hospital of Anhui Medical University, Hefei, Anhui, 230601, China
| | - Xiaoping Li
- Department of Chinese Medicine, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, 450052, China
| | - Yong Wang
- Department of General Surgery, The Second Affiliated Hospital of Anhui Medical University, Hefei, Anhui, 230601, China.
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15
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Gou H, Zhao M, Xu H, Yuan J, He W, Zhu M, Ding H, Yi L, Chen J. CSFV induced mitochondrial fission and mitophagy to inhibit apoptosis. Oncotarget 2018; 8:39382-39400. [PMID: 28455958 PMCID: PMC5503620 DOI: 10.18632/oncotarget.17030] [Citation(s) in RCA: 49] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/17/2016] [Accepted: 03/17/2017] [Indexed: 01/01/2023] Open
Abstract
Classical swine fever virus (CSFV), which causes typical clinical characteristics in piglets, including hemorrhagic syndrome and immunosuppression, is linked to hepatitis C and dengue virus. Oxidative stress and a reduced mitochondrial transmembrane potential are disturbed in CSFV-infected cells. The balance of mitochondrial dynamics is essential for cellular homeostasis. In this study, we offer the first evidence that CSFV induces mitochondrial fission and mitophagy to inhibit host cell apoptosis for persistent infection. The formation of mitophagosomes and decline in mitochondrial mass relevant to mitophagy were detected in CSFV-infected cells. CSFV infection increased the expression and mitochondrial translocation of Pink and Parkin. Upon activation of the PINK1 and Parkin pathways, Mitofusin 2 (MFN2), a mitochondrial fusion mediator, was ubiquitinated and degraded in CSFV-infected cells. Mitophagosomes and mitophagolysosomes induced by CSFV were, respectively, observed by the colocalization of LC3-associated mitochondria with Parkin or lysosomes. In addition, a sensitive dual fluorescence reporter (mito-mRFP-EGFP) was utilized to analyze the delivery of mitophagosomes to lysosomes. Mitochondrial fission caused by CSFV infection was further determined by mitochondrial fragmentation and Drp1 translocation into mitochondria using a confocal microscope. The preservation of mitochondrial proteins, upregulated apoptotic signals and decline of viral replication resulting from the silencing of Drp1 and Parkin in CSFV-infected cells suggested that CSFV induced mitochondrial fission and mitophagy to enhance cell survival and viral persistence. Our data for mitochondrial fission and selective mitophagy in CSFV-infected cells reveal a unique view of the pathogenesis of CSFV infection and provide new avenues for the development of antiviral strategies.
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Affiliation(s)
- Hongchao Gou
- College of Veterinary Medicine, South China Agricultural University, Guangzhou, People's Republic of China
| | - Mingqiu Zhao
- College of Veterinary Medicine, South China Agricultural University, Guangzhou, People's Republic of China
| | - Hailuan Xu
- College of Veterinary Medicine, South China Agricultural University, Guangzhou, People's Republic of China
| | - Jin Yuan
- College of Veterinary Medicine, South China Agricultural University, Guangzhou, People's Republic of China
| | - Wencheng He
- College of Veterinary Medicine, South China Agricultural University, Guangzhou, People's Republic of China
| | - Mengjiao Zhu
- College of Veterinary Medicine, South China Agricultural University, Guangzhou, People's Republic of China
| | - Hongxing Ding
- College of Veterinary Medicine, South China Agricultural University, Guangzhou, People's Republic of China
| | - Lin Yi
- College of Veterinary Medicine, South China Agricultural University, Guangzhou, People's Republic of China
| | - Jinding Chen
- College of Veterinary Medicine, South China Agricultural University, Guangzhou, People's Republic of China
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16
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Scrima R, Piccoli C, Moradpour D, Capitanio N. Targeting Endoplasmic Reticulum and/or Mitochondrial Ca 2+ Fluxes as Therapeutic Strategy for HCV Infection. Front Chem 2018; 6:73. [PMID: 29619366 PMCID: PMC5871704 DOI: 10.3389/fchem.2018.00073] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/26/2018] [Accepted: 03/06/2018] [Indexed: 01/16/2023] Open
Abstract
Chronic hepatitis C is characterized by metabolic disorders and by a microenvironment in the liver dominated by oxidative stress, inflammation and regeneration processes that can in the long term lead to liver cirrhosis and hepatocellular carcinoma. Several lines of evidence suggest that mitochondrial dysfunctions play a central role in these processes. However, how these dysfunctions are induced by the virus and whether they play a role in disease progression and neoplastic transformation remains to be determined. Most in vitro studies performed so far have shown that several of the hepatitis C virus (HCV) proteins also localize to mitochondria, but the consequences of these interactions on mitochondrial functions remain contradictory and need to be confirmed in the context of productively replicating virus and physiologically relevant in vitro and in vivo model systems. In the past decade we have been proposing a temporal sequence of events in the HCV-infected cell whereby the primary alteration is localized at the mitochondria-associated ER membranes and causes release of Ca2+ from the ER, followed by uptake into mitochondria. This ensues successive mitochondrial dysfunction leading to the generation of reactive oxygen and nitrogen species and a progressive metabolic adaptive response consisting in decreased oxidative phosphorylation and enhanced aerobic glycolysis and lipogenesis. Here we resume the major results provided by our group in the context of HCV-mediated alterations of the cellular inter-compartmental calcium flux homeostasis and present new evidence suggesting targeting of ER and/or mitochondrial calcium transporters as a novel therapeutic strategy.
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Affiliation(s)
- Rosella Scrima
- Department of Clinical and Experimental Medicine, University of Foggia, Foggia, Italy
| | - Claudia Piccoli
- Department of Clinical and Experimental Medicine, University of Foggia, Foggia, Italy
| | - Darius Moradpour
- Service of Gastroenterology and Hepatology, Centre Hospitalier Universitaire Vaudois, University of Lausanne, Lausanne, Switzerland
| | - Nazzareno Capitanio
- Department of Clinical and Experimental Medicine, University of Foggia, Foggia, Italy
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17
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Mitochondria-associated membranes (MAMs) and inflammation. Cell Death Dis 2018; 9:329. [PMID: 29491386 PMCID: PMC5832426 DOI: 10.1038/s41419-017-0027-2] [Citation(s) in RCA: 229] [Impact Index Per Article: 32.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/03/2017] [Revised: 09/26/2017] [Accepted: 09/27/2017] [Indexed: 12/18/2022]
Abstract
The endoplasmic reticulum (ER) and mitochondria are tightly associated with very dynamic platforms termed mitochondria-associated membranes (MAMs). MAMs provide an excellent scaffold for crosstalk between the ER and mitochondria and play a pivotal role in different signaling pathways that allow rapid exchange of biological molecules to maintain cellular health. However, dysfunctions in the ER–mitochondria architecture are associated with pathological conditions and human diseases. Inflammation has emerged as one of the various pathways that MAMs control. Inflammasome components and other inflammatory factors promote the release of pro-inflammatory cytokines that sustain pathological conditions. In this review, we summarize the critical role of MAMs in initiating inflammation in the cellular defense against pathogenic infections and the association of MAMs with inflammation-mediated diseases.
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18
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Oxidative stress, a trigger of hepatitis C and B virus-induced liver carcinogenesis. Oncotarget 2018; 8:3895-3932. [PMID: 27965466 PMCID: PMC5354803 DOI: 10.18632/oncotarget.13904] [Citation(s) in RCA: 120] [Impact Index Per Article: 17.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/22/2016] [Accepted: 12/05/2016] [Indexed: 12/11/2022] Open
Abstract
Virally induced liver cancer usually evolves over long periods of time in the context of a strongly oxidative microenvironment, characterized by chronic liver inflammation and regeneration processes. They ultimately lead to oncogenic mutations in many cellular signaling cascades that drive cell growth and proliferation. Oxidative stress, induced by hepatitis viruses, therefore is one of the factors that drives the neoplastic transformation process in the liver. This review summarizes current knowledge on oxidative stress and oxidative stress responses induced by human hepatitis B and C viruses. It focuses on the molecular mechanisms by which these viruses activate cellular enzymes/systems that generate or scavenge reactive oxygen species (ROS) and control cellular redox homeostasis. The impact of an altered cellular redox homeostasis on the initiation and establishment of chronic viral infection, as well as on the course and outcome of liver fibrosis and hepatocarcinogenesis will be discussed The review neither discusses reactive nitrogen species, although their metabolism is interferes with that of ROS, nor antioxidants as potential therapeutic remedies against viral infections, both subjects meriting an independent review.
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19
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Cavallari I, Scattolin G, Silic-Benussi M, Raimondi V, D'Agostino DM, Ciminale V. Mitochondrial Proteins Coded by Human Tumor Viruses. Front Microbiol 2018; 9:81. [PMID: 29467726 PMCID: PMC5808139 DOI: 10.3389/fmicb.2018.00081] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2017] [Accepted: 01/12/2018] [Indexed: 12/26/2022] Open
Abstract
Viruses must exploit the cellular biosynthetic machinery and evade cellular defense systems to complete their life cycles. Due to their crucial roles in cellular bioenergetics, apoptosis, innate immunity and redox balance, mitochondria are important functional targets of many viruses, including tumor viruses. The present review describes the interactions between mitochondria and proteins coded by the human tumor viruses human T-cell leukemia virus type 1, Epstein-Barr virus, Kaposi's sarcoma-associated herpesvirus, human hepatitis viruses B and C, and human papillomavirus, and highlights how these interactions contribute to viral replication, persistence and transformation.
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Affiliation(s)
| | - Gloria Scattolin
- Department of Surgery, Oncology, and Gastroenterology, University of Padova, Padova, Italy
| | | | | | | | - Vincenzo Ciminale
- Veneto Institute of Oncology IOV-IRRCS, Padova, Italy.,Department of Surgery, Oncology, and Gastroenterology, University of Padova, Padova, Italy
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20
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Abstract
Viruses use synthetic mechanism and organelles of the host cells to facilitate their replication and make new viruses. Host's ATP provides necessary energy. Hepatitis C virus (HCV) is a major cause of liver disease. Like other positive-strand RNA viruses, the HCV genome is thought to be synthesized by the replication complex, which consists of viral- and host cell-derived factors, in tight association with structurally rearranged vesicle-like cytoplasmic membranes. The virus-induced remodeling of subcellular membranes, which protect the viral RNA from nucleases in the cytoplasm, promotes efficient replication of HCV genome. The assembly of HCV particle involves interactions between viral structural and nonstructural proteins and pathways related to lipid metabolisms in a concerted fashion. Association of viral core protein, which forms the capsid, with lipid droplets appears to be a prerequisite for early steps of the assembly, which are closely linked with the viral genome replication. This review presents the recent progress in understanding the mechanisms for replication and assembly of HCV through its interactions with organelles or distinct organelle-like structures.
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Affiliation(s)
- Tetsuro Suzuki
- Department of Virology and Parasitology, Hamamatsu University School of Medicine, Hamamatsu, Shizuoka, 431-3192, Japan.
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21
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Cao L, Quan XB, Zeng WJ, Yang XO, Wang MJ. Mechanism of Hepatocyte Apoptosis. J Cell Death 2016; 9:19-29. [PMID: 28058033 PMCID: PMC5201115 DOI: 10.4137/jcd.s39824] [Citation(s) in RCA: 62] [Impact Index Per Article: 6.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/01/2016] [Revised: 08/14/2016] [Accepted: 08/18/2016] [Indexed: 12/12/2022] Open
Abstract
Hepatocyte apoptosis plays important roles in both the removal of external microorganisms and the occurrence and development of liver diseases. Different conditions, such as virus infection, fatty liver disease, hepatic ischemia reperfusion, and drug-induced liver injury, are accompanied by hepatocyte apoptosis. This review summarizes recent research on the mechanism of hepatocyte apoptosis involving the classical extrinsic and intrinsic apoptotic pathways, endoplasmic reticulum stress, and oxidative stress-induced apoptosis. We emphasized the major causes of apoptosis according to the characteristics of different liver diseases. Several concerns regarding future research and clinical application are also raised.
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Affiliation(s)
- Lei Cao
- Research Center on Aging and Medicine, Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Fudan University, Shanghai, China
| | - Xi-Bing Quan
- Research Center on Aging and Medicine, Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Fudan University, Shanghai, China
| | - Wen-Jiao Zeng
- Department of Pathology, School of Basic Medical Sciences, Fudan University, Shanghai, China
| | - Xiao-Ou Yang
- Department of Gastroenterology, Peking Union Medical College Hospital, Beijing, China
| | - Ming-Jie Wang
- Research Center on Aging and Medicine, Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Fudan University, Shanghai, China
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22
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Medvedev R, Hildt E, Ploen D. Look who's talking-the crosstalk between oxidative stress and autophagy supports exosomal-dependent release of HCV particles. Cell Biol Toxicol 2016; 33:211-231. [PMID: 27987184 DOI: 10.1007/s10565-016-9376-3] [Citation(s) in RCA: 31] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/27/2016] [Accepted: 12/04/2016] [Indexed: 12/12/2022]
Abstract
Autophagy is a highly conserved and regulated intracellular lysosomal degradation pathway that is essential for cell survival. Dysregulation has been linked to the development of various human diseases, including neurodegeneration and tumorigenesis, infection, and aging. Besides, many viruses hijack the autophagosomal pathway to support their life cycle. The hepatitis C virus (HCV), a major cause of chronic liver diseases worldwide, has been described to induce autophagy. The autophagosomal pathway can be further activated in response to elevated levels of reactive oxygen species (ROS). HCV impairs the Nrf2/ARE-dependent induction of ROS-detoxifying enzymes by a so far unprecedented mechanism. In line with this, this review aims to discuss the relevance of HCV-dependent elevated ROS levels for the induction of autophagy as a result of the impaired Nrf2 signaling and the described crosstalk between p62 and the Nrf2/Keap1 signaling pathway. Moreover, autophagy is functionally connected to the endocytic pathway as components of the endosomal trafficking are involved in the maturation of autophagosomes. The release of HCV particles is still not fully understood. Recent studies suggest an involvement of exosomes that originate from the endosomal pathway in viral release. In line with this, it is tempting to speculate whether HCV-dependent elevated ROS levels induce autophagy to support exosome-mediated release of viral particles. Based on recent findings, in this review, we will further highlight the impact of HCV-induced autophagy and its interplay with the endosomal pathway as a novel mechanism for the release of HCV particles.
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Affiliation(s)
- Regina Medvedev
- Department of Virology, Paul-Ehrlich-Institut, Paul-Ehrlich-Straße 51-59, 63225, Langen, Germany
| | - Eberhard Hildt
- Department of Virology, Paul-Ehrlich-Institut, Paul-Ehrlich-Straße 51-59, 63225, Langen, Germany.,Deutsches Zentrum für Infektionsforschung (DZIF), Gießen, Marburg, Langen, Germany
| | - Daniela Ploen
- Department of Virology, Paul-Ehrlich-Institut, Paul-Ehrlich-Straße 51-59, 63225, Langen, Germany.
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23
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Dengue virus induces mitochondrial elongation through impairment of Drp1-triggered mitochondrial fission. Virology 2016; 500:149-160. [PMID: 27816895 DOI: 10.1016/j.virol.2016.10.022] [Citation(s) in RCA: 54] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/09/2016] [Revised: 10/18/2016] [Accepted: 10/24/2016] [Indexed: 02/08/2023]
Abstract
Mitochondria are highly dynamic organelles that undergo continuous cycles of fission and fusion to maintain essential cellular functions. An imbalance between these two processes can result in many pathophysiological outcomes. Dengue virus (DENV) interacts with cellular organelles, including mitochondria, to successfully replicate in cells. This study used live-cell imaging and found an increase in mitochondrial length and respiration during DENV infection. The level of mitochondrial fission protein, Dynamin-related protein 1 (Drp1), was decreased on mitochondria during DENV infection, as well as Drp1 phosphorylated on serine 616, which is important for mitochondrial fission. DENV proteins NS4b and NS3 were also associated with subcellular fractions of mitochondria. Induction of fission through uncoupling of mitochondria or overexpression of Drp1 wild-type and Drp1 with a phosphomimetic mutation (S616D) significantly reduced viral replication. These results demonstrate that DENV infection causes an imbalance in mitochondrial dynamics by inhibiting Drp1-triggered mitochondrial fission, which promotes viral replication.
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24
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Meyers NL, Fontaine KA, Kumar GR, Ott M. Entangled in a membranous web: ER and lipid droplet reorganization during hepatitis C virus infection. Curr Opin Cell Biol 2016; 41:117-24. [PMID: 27240021 DOI: 10.1016/j.ceb.2016.05.003] [Citation(s) in RCA: 28] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/09/2016] [Revised: 05/04/2016] [Accepted: 05/05/2016] [Indexed: 12/19/2022]
Abstract
Hepatitis C virus (HCV) is a major cause of liver disease worldwide. To establish and maintain chronic infection, HCV extensively rearranges cellular organelles to generate distinct compartments for viral RNA replication and virion assembly. Here, we review our current knowledge of how HCV proliferates and remodels ER-derived membranes while preserving and expanding associated lipid droplets during viral infection. Unraveling the molecular mechanisms responsible for HCV-induced membrane reorganization will enhance our understanding of the HCV life-cycle, the associated liver pathology, and the biology of the ER:lipid droplet interface in general.
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Affiliation(s)
- Nathan L Meyers
- Gladstone Institutes, University of California San Francisco, 1650 Owens Street, San Francisco, CA 94941, United States
| | - Krystal A Fontaine
- Gladstone Institutes, University of California San Francisco, 1650 Owens Street, San Francisco, CA 94941, United States
| | - G Renuka Kumar
- Gladstone Institutes, University of California San Francisco, 1650 Owens Street, San Francisco, CA 94941, United States
| | - Melanie Ott
- Gladstone Institutes, University of California San Francisco, 1650 Owens Street, San Francisco, CA 94941, United States.
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25
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Wang SR, Min YQ, Wang JQ, Liu CX, Fu BS, Wu F, Wu LY, Qiao ZX, Song YY, Xu GH, Wu ZG, Huang G, Peng NF, Huang R, Mao WX, Peng S, Chen YQ, Zhu Y, Tian T, Zhang XL, Zhou X. A highly conserved G-rich consensus sequence in hepatitis C virus core gene represents a new anti-hepatitis C target. SCIENCE ADVANCES 2016; 2:e1501535. [PMID: 27051880 PMCID: PMC4820367 DOI: 10.1126/sciadv.1501535] [Citation(s) in RCA: 101] [Impact Index Per Article: 11.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/29/2015] [Accepted: 02/18/2016] [Indexed: 05/24/2023]
Abstract
G-quadruplex (G4) is one of the most important secondary structures in nucleic acids. Until recently, G4 RNAs have not been reported in any ribovirus, such as the hepatitis C virus. Our bioinformatics analysis reveals highly conserved guanine-rich consensus sequences within the core gene of hepatitis C despite the high genetic variability of this ribovirus; we further show using various methods that such consensus sequences can fold into unimolecular G4 RNA structures, both in vitro and under physiological conditions. Furthermore, we provide direct evidences that small molecules specifically targeting G4 can stabilize this structure to reduce RNA replication and inhibit protein translation of intracellular hepatitis C. Ultimately, the stabilization of G4 RNA in the genome of hepatitis C represents a promising new strategy for anti-hepatitis C drug development.
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Affiliation(s)
- Shao-Ru Wang
- College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan 430072, Hubei, China
| | - Yuan-Qin Min
- State Key Laboratory of Virology and Hubei Province Key Laboratory of Allergy and Immunology and Department of Immunology, School of Medicine, Wuhan University, Wuhan 430071, Hubei, China
| | - Jia-Qi Wang
- College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan 430072, Hubei, China
| | - Chao-Xing Liu
- College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan 430072, Hubei, China
| | - Bo-Shi Fu
- College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan 430072, Hubei, China
| | - Fan Wu
- College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan 430072, Hubei, China
| | - Ling-Yu Wu
- College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan 430072, Hubei, China
| | - Zhi-Xian Qiao
- Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, Hubei, China
| | - Yan-Yan Song
- College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan 430072, Hubei, China
| | - Guo-Hua Xu
- Wuhan Institute of Physics and Mathematics, Chinese Academy of Sciences, Wuhan 430071, Hubei, China
| | - Zhi-Guo Wu
- College of Life Sciences, Wuhan University, Wuhan 430072, Hubei, China
| | - Gai Huang
- College of Life Sciences, Wuhan University, Wuhan 430072, Hubei, China
| | - Nan-Fang Peng
- College of Life Sciences, Wuhan University, Wuhan 430072, Hubei, China
| | - Rong Huang
- College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan 430072, Hubei, China
| | - Wu-Xiang Mao
- College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan 430072, Hubei, China
| | - Shuang Peng
- College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan 430072, Hubei, China
| | - Yu-Qi Chen
- College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan 430072, Hubei, China
| | - Ying Zhu
- College of Life Sciences, Wuhan University, Wuhan 430072, Hubei, China
| | - Tian Tian
- College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan 430072, Hubei, China
| | - Xiao-Lian Zhang
- State Key Laboratory of Virology and Hubei Province Key Laboratory of Allergy and Immunology and Department of Immunology, School of Medicine, Wuhan University, Wuhan 430071, Hubei, China
| | - Xiang Zhou
- College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Wuhan 430072, Hubei, China
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Poortahmasebi V, Poorebrahim M, Najafi S, Jazayeri SM, Alavian SM, Arab SS, Ghavami S, Alavian SE, Rezaei Moghadam A, Amiri M. How Hepatitis C Virus Leads to Hepatocellular Carcinoma: A Network-Based Study. HEPATITIS MONTHLY 2016; 16:e36005. [PMID: 27148389 PMCID: PMC4852094 DOI: 10.5812/hepatmon.36005] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 01/02/2016] [Accepted: 01/20/2016] [Indexed: 12/11/2022]
Abstract
BACKGROUND Hepatitis C virus (HCV) has been known as a major cause of hepatocellular carcinoma (HCC) worldwide. However, the distinct molecular mechanisms underlying the effects of HCV proteins on the HCC progression have remained unclear. OBJECTIVES In the present study, we studied the possible role of HCV in the HCC initiation and invasion using topological analysis of protein-protein interaction (PPI) networks. MATERIALS AND METHODS After analysis with GEO2R, a PPI network of differentially expressed genes (DEGs) was constructed for both chronic HCV and HCC samples. The STRING and GeneMANIA databases were used to determine the putative interactions between DEGs. In parallel, the functional annotation of DEGs was performed using g: Profiler web tool. The topological analysis and network visualization was carried outperformed using Cytoscape software and the top hub genes were identified. We determined the hub genes-related miRNAs using miRTarBase server and reconstructed a miRNA-Hubgene network. RESULTS Based on the topological analysis of miRNA-Hubgene network, we identified the key hub miRNAs. In order to identify the most important common sub-network, we aligned two PPI networks using NETAL tool. The c-Jun gene was identified as the most important hub gene in both HCV and HCC networks. Furthermore, the hsa-miR-34a, hsa-miR-155, hsa-miR-24, hsa-miR-744 and hsa-miR-92a were recognized as the most important hub miRNAs with positive correlation in the chronic HCV and HCC samples. Functional annotation of differentially expressed miRNAs (DEMs) using the tool for annotations of human miRNAs (TAM) revealed that there is a considerable overlap between miRNA gene expression profiles of HCV-infected and HCC cells. CONCLUSIONS Our results revealed the possible crucial genes and miRNAs involved in the initiation and progression of HCC cells infected with HCV.
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Affiliation(s)
- Vahdat Poortahmasebi
- Hepatitis B Molecular Laboratory, Department of Virology, School of Public Health, Tehran University of Medical Sciences, Tehran, IR Iran
| | - Mansour Poorebrahim
- Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran, IR Iran
- Corresponding Author: Mansour Poorebrahim, Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran, IR Iran. Tel: +98-9120192664, E-mail:
| | - Saeideh Najafi
- Department of Microbiology, Tonekabon Branch, Islamic Azad University, Tonekabon, IR Iran
| | - Seyed Mohammad Jazayeri
- Hepatitis B Molecular Laboratory, Department of Virology, School of Public Health, Tehran University of Medical Sciences, Tehran, IR Iran
| | | | - Seyed Shahriar Arab
- Department of Biophysics, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, IR Iran
| | - Saeid Ghavami
- Department of Human Anatomy and Cell Science, University of Manitoba, Winnipeg, Canada
| | | | - Adel Rezaei Moghadam
- Department of Human Anatomy and Cell Science, University of Manitoba, Winnipeg, Canada
| | - Mehdi Amiri
- Department of Cell Biology and Anatomy, Schulich School of Medicine and Dentistry, Western University, London, Canada
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27
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HCV and Oxidative Stress: Implications for HCV Life Cycle and HCV-Associated Pathogenesis. OXIDATIVE MEDICINE AND CELLULAR LONGEVITY 2016; 2016:9012580. [PMID: 26955431 PMCID: PMC4756209 DOI: 10.1155/2016/9012580] [Citation(s) in RCA: 54] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 11/25/2015] [Accepted: 01/14/2016] [Indexed: 12/15/2022]
Abstract
HCV (hepatitis C virus) is a member of the Flaviviridae family that contains a single-stranded positive-sense RNA genome of approximately 9600 bases. HCV is a major causative agent for chronic liver diseases such as steatosis, fibrosis, cirrhosis, and hepatocellular carcinoma which are caused by multifactorial processes. Elevated levels of reactive oxygen species (ROS) are considered as a major factor contributing to HCV-associated pathogenesis. This review summarizes the mechanisms involved in formation of ROS in HCV replicating cells and describes the interference of HCV with ROS detoxifying systems. The relevance of ROS for HCV-associated pathogenesis is reviewed with a focus on the interference of elevated ROS levels with processes controlling liver regeneration. The overview about the impact of ROS for the viral life cycle is focused on the relevance of autophagy for the HCV life cycle and the crosstalk between HCV, elevated ROS levels, and the induction of autophagy.
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28
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Ramage HR, Kumar GR, Verschueren E, Johnson JR, Von Dollen J, Johnson T, Newton B, Shah P, Horner J, Krogan NJ, Ott M. A combined proteomics/genomics approach links hepatitis C virus infection with nonsense-mediated mRNA decay. Mol Cell 2015; 57:329-340. [PMID: 25616068 DOI: 10.1016/j.molcel.2014.12.028] [Citation(s) in RCA: 114] [Impact Index Per Article: 11.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/09/2014] [Revised: 10/28/2014] [Accepted: 12/16/2014] [Indexed: 12/22/2022]
Abstract
Hepatitis C virus (HCV) is a leading cause of liver disease, but insight into virus-host interactions remains limited. We systematically used affinity purification/mass spectrometry to define the host interactions of all ten HCV proteins in hepatoma cells. We combined these studies with RNAi knockdown of corresponding genes using a two-step scoring approach to generate a map of 139 high-confidence HCV-host protein-protein interactions. We found mitochondrial proteins highly involved in HCV infection and characterized an interaction between the viral core protein and host protein within bgcn homolog (WIBG). Expression of core prevents WIBG from binding its regular interaction partners Y14 and Magoh, two known mediators of the nonsense-mediated mRNA decay pathway. We discovered that this surveillance pathway is disrupted in HCV-infected cells, causing potentially harmful transcripts to accumulate. Our study provides a comprehensive view of HCV-host interactions and uncovers mechanisms for how HCV perturbs host functions during infection.
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Affiliation(s)
- Holly R Ramage
- Gladstone Institutes, 1650 Owens Street, San Francisco, CA 94158, USA
- University of California, San Francisco, San Francisco, CA 94158, USA
| | - G Renuka Kumar
- Gladstone Institutes, 1650 Owens Street, San Francisco, CA 94158, USA
- University of California, San Francisco, San Francisco, CA 94158, USA
| | - Erik Verschueren
- University of California, San Francisco, San Francisco, CA 94158, USA
- QB3, California Institute for Quantitative Biosciences, San Francisco, CA 94158, USA
| | - Jeffrey R Johnson
- Gladstone Institutes, 1650 Owens Street, San Francisco, CA 94158, USA
- University of California, San Francisco, San Francisco, CA 94158, USA
| | - John Von Dollen
- University of California, San Francisco, San Francisco, CA 94158, USA
- QB3, California Institute for Quantitative Biosciences, San Francisco, CA 94158, USA
| | - Tasha Johnson
- University of California, San Francisco, San Francisco, CA 94158, USA
- QB3, California Institute for Quantitative Biosciences, San Francisco, CA 94158, USA
| | - Billy Newton
- University of California, San Francisco, San Francisco, CA 94158, USA
- QB3, California Institute for Quantitative Biosciences, San Francisco, CA 94158, USA
| | - Priya Shah
- University of California, San Francisco, San Francisco, CA 94158, USA
- QB3, California Institute for Quantitative Biosciences, San Francisco, CA 94158, USA
| | - Julie Horner
- Thermo Fisher Scientific, 355 River Oaks Pkwy, San Jose, CA 95134, USA
| | - Nevan J Krogan
- Gladstone Institutes, 1650 Owens Street, San Francisco, CA 94158, USA
- University of California, San Francisco, San Francisco, CA 94158, USA
- QB3, California Institute for Quantitative Biosciences, San Francisco, CA 94158, USA
| | - Melanie Ott
- Gladstone Institutes, 1650 Owens Street, San Francisco, CA 94158, USA
- University of California, San Francisco, San Francisco, CA 94158, USA
- Liver Center, University of California, San Francisco, CA 94143, USA
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29
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Cellular stress responses in hepatitis C virus infection: Mastering a two-edged sword. Virus Res 2015; 209:100-17. [PMID: 25836277 DOI: 10.1016/j.virusres.2015.03.013] [Citation(s) in RCA: 34] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2014] [Revised: 03/21/2015] [Accepted: 03/23/2015] [Indexed: 12/11/2022]
Abstract
Hepatitis C virus (HCV) infection affects chronically more than 150 million humans worldwide. Chronic HCV infection causes severe liver disease and hepatocellular carcinoma. While immune response-mediated events are major players in HCV pathogenesis, the impact that viral replication has on cellular homeostasis is increasingly recognized as a necessary contributor to pathological manifestations of HCV infection such as steatosis, insulin-resistance or liver cancer. In this review, we will briefly overview the different cellular stress pathways that are induced by hepatitis C virus infection, the response that the cell promotes to attempt regaining homeostasis or to induce dysfunctional cell death, and how the virus co-opts these response mechanisms to promote both viral replication and survival of the infected cell. We will review the role of unfolded protein and oxidative stress responses as well as the role of auto- and mitophagy in HCV infection. Finally, we will discuss the recent discovery of a cellular chaperone involved in stress responses, the sigma-1 receptor, as a cellular factor required at the onset of HCV infection and the potential molecular events underlying the proviral role of this cellular factor in HCV infection.
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30
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Douglas DN, Kneteman NM. Generation of improved mouse models for the study of hepatitis C virus. Eur J Pharmacol 2015; 759:313-25. [PMID: 25814250 DOI: 10.1016/j.ejphar.2015.03.022] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/19/2015] [Revised: 03/06/2015] [Accepted: 03/12/2015] [Indexed: 12/15/2022]
Abstract
Approximately 3% of the world׳s population suffers from chronic infections with hepatitis C virus (HCV). Although current treatment regimes are capable of effectively eradicating HCV infection from these patients, the cost of these combinations of direct-acting antivirals are prohibitive. Approximately 80% of untreated chronic HCV carriers will be at high risk for developing severe liver disease, including fibrosis, cirrhosis, and hepatocellular carcinoma. A vaccine is urgently needed to lessen this global burden. Besides humans, HCV infection can be experimentally transmitted to chimpanzees, and this is the best model for studies of HCV infection and related innate and adaptive immune responses. Although the chimpanzee model yielded valuable insight, limited availability, high cost and ethical considerations limit their utility. The only small animal models of robust HCV infection are highly immunodeficient mice with human chimeric livers. However, these mice cannot be used to study adaptive immune responses and therefore a more relevant animal model is needed to assist in vaccine development. Novel strains of immunodeficient mice have been developed that allow for the engraftment of human hepatopoietic stem cells, as well as functional human lymphoid cells and tissues, effectively creating human immune systems in otherwise immunodeficient mice. These humanized mice are rapidly emerging as pre-clinical bridges for numerous pathogens that, like HCV, only cause infectious disease in humans. This review highlights the potential these new models have for changing the current landscape for HCV research and vaccine development.
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Affiliation(s)
- Donna N Douglas
- Department of Surgery, University of Alberta, Edmonton, Alberta, Canada T6G 2E1; Li Ka Shing Institute of Virology, University of Alberta, Edmonton, Alberta, Canada T6G 2E1.
| | - Norman M Kneteman
- Department of Surgery, University of Alberta, Edmonton, Alberta, Canada T6G 2E1; Li Ka Shing Institute of Virology, University of Alberta, Edmonton, Alberta, Canada T6G 2E1; KMT Hepatech Inc., Edmonton, Alberta, Canada T6G 2M9
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31
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Horner SM, Wilkins C, Badil S, Iskarpatyoti J, Gale M. Proteomic analysis of mitochondrial-associated ER membranes (MAM) during RNA virus infection reveals dynamic changes in protein and organelle trafficking. PLoS One 2015; 10:e0117963. [PMID: 25734423 PMCID: PMC4348417 DOI: 10.1371/journal.pone.0117963] [Citation(s) in RCA: 87] [Impact Index Per Article: 8.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/20/2014] [Accepted: 01/06/2015] [Indexed: 02/07/2023] Open
Abstract
RIG-I pathway signaling of innate immunity against RNA virus infection is organized between the ER and mitochondria on a subdomain of the ER called the mitochondrial-associated ER membrane (MAM). The RIG-I adaptor protein MAVS transmits downstream signaling of antiviral immunity, with signaling complexes assembling on the MAM in association with mitochondria and peroxisomes. To identify components that regulate MAVS signalosome assembly on the MAM, we characterized the proteome of MAM, ER, and cytosol from cells infected with either chronic (hepatitis C) or acute (Sendai) RNA virus infections, as well as mock-infected cells. Comparative analysis of protein trafficking dynamics during both chronic and acute viral infection reveals differential protein profiles in the MAM during RIG-I pathway activation. We identified proteins and biochemical pathways recruited into and out of the MAM in both chronic and acute RNA viral infections, representing proteins that drive immunity and/or regulate viral replication. In addition, by using this comparative proteomics approach, we identified 3 new MAVS-interacting proteins, RAB1B, VTN, and LONP1, and defined LONP1 as a positive regulator of the RIG-I pathway. Our proteomic analysis also reveals a dynamic cross-talk between subcellular compartments during both acute and chronic RNA virus infection, and demonstrates the importance of the MAM as a central platform that coordinates innate immune signaling to initiate immunity against RNA virus infection.
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Affiliation(s)
- Stacy M. Horner
- Department of Immunology, University of Washington, Seattle, Washington, United States of America
- Department of Medicine, Duke University Medical Center, Durham, North Carolina, United States of America
| | - Courtney Wilkins
- Department of Immunology, University of Washington, Seattle, Washington, United States of America
| | - Samantha Badil
- Department of Immunology, University of Washington, Seattle, Washington, United States of America
| | - Jason Iskarpatyoti
- Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina, United States of America
| | - Michael Gale
- Department of Immunology, University of Washington, Seattle, Washington, United States of America
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32
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Bassiony MM, Yousef A, Raya Y, El-Shabrawi A, Fouad E, El-Shafeey M. Cognitive impairment in relation to depression, anxiety and virological response in hepatitis C patients in Egypt. Int J Psychiatry Clin Pract 2015; 19:208-15. [PMID: 26099337 DOI: 10.3109/13651501.2015.1064964] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/18/2023]
Abstract
OBJECTIVE Cognitive impairment commonly occurs in hepatitis C virus (HCV) patients. The objective of this study was to estimate the prevalence and sociodemographic and clinical correlates of cognitive impairment in HCV patients before and after 12 weeks of interferon treatment in comparison with a control group. METHODS Hundred and sixteen consecutive HCV patients were included in the study and divided into treated and untreated groups. All patients were assessed using sociodemographic and clinical questionnaire, Montreal Cognitive Assessment Scale (MOCA) and Hospital Anxiety and Depression Scale (HADS) before and after 12 weeks of treatment or observation. RESULTS Thirty-eight percent of treated patients showed cognitive impairment at baseline, which increased to 69% after 12 weeks of interferon treatment. This cognitive impairment was reflected in the total MOCA score and in visuo-constructional skills, attention, concentration, working memory, language, and short-term memory, which was not shown by untreated group after 12 weeks of observation. Cognitive impairment was associated with low education, but not with depression, anxiety, or virological response. CONCLUSIONS Cognitive impairment is common in HCV patients and increases significantly after interferon treatment. It is not related to depression or anxiety in HCV patients. Future research should focus on prevention, treatment and outcome of cognitive impairment in HCV patients, particularly those receiving interferon therapy.
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Affiliation(s)
- M M Bassiony
- a Department of Psychiatry , Faculty of Medicine, Zagazig University , Egypt
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33
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Cheng ML, Weng SF, Kuo CH, Ho HY. Enterovirus 71 induces mitochondrial reactive oxygen species generation that is required for efficient replication. PLoS One 2014; 9:e113234. [PMID: 25401329 PMCID: PMC4234665 DOI: 10.1371/journal.pone.0113234] [Citation(s) in RCA: 62] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/14/2014] [Accepted: 10/22/2014] [Indexed: 12/16/2022] Open
Abstract
Redox homeostasis is an important host factor determining the outcome of infectious disease. Enterovirus 71 (EV71) infection has become an important endemic disease in Southeast Asia and China. We have previously shown that oxidative stress promotes viral replication, and progeny virus induces oxidative stress in host cells. The detailed mechanism for reactive oxygen species (ROS) generation in infected cells remains elusive. In the current study, we demonstrate that mitochondria were a major ROS source in EV71-infected cells. Mitochondria in productively infected cells underwent morphologic changes and exhibited functional anomalies, such as a decrease in mitochondrial electrochemical potential ΔΨm and an increase in oligomycin-insensitive oxygen consumption. Respiratory control ratio of mitochondria from infected cells was significantly lower than that of normal cells. The total adenine nucleotide pool and ATP content of EV71-infected cells significantly diminished. However, there appeared to be a compensatory increase in mitochondrial mass. Treatment with mito-TEMPO reduced eIF2α phosphorylation and viral replication, suggesting that mitochondrial ROS act to promote viral replication. It is plausible that EV71 infection induces mitochondrial ROS generation, which is essential to viral replication, at the sacrifice of efficient energy production, and that infected cells up-regulate biogenesis of mitochondria to compensate for their functional defect.
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Affiliation(s)
- Mei-Ling Cheng
- Department of Biomedical Sciences, College of Medicine, Chang Gung University, Tao-Yuan, Taiwan
- Healthy Aging Research Center, Chang Gung University, Tao-Yuan, Taiwan
- Metabolomics Core Laboratory, Chang Gung University, Tao-Yuan, Taiwan
| | - Shiue-Fen Weng
- Department of Medical Biotechnology and Laboratory Science, College of Medicine, Chang Gung University, Tao-Yuan, Taiwan
| | - Chih-Hao Kuo
- Department of Medical Biotechnology and Laboratory Science, College of Medicine, Chang Gung University, Tao-Yuan, Taiwan
| | - Hung-Yao Ho
- Healthy Aging Research Center, Chang Gung University, Tao-Yuan, Taiwan
- Department of Medical Biotechnology and Laboratory Science, College of Medicine, Chang Gung University, Tao-Yuan, Taiwan
- Office of Research and Development, Chang Gung Memorial Hospital, Tao-Yuan, Taiwan
- * E-mail:
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34
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Choi YB, Harhaj EW. Functional implications of mitochondrial reactive oxygen species generated by oncogenic viruses. ACTA ACUST UNITED AC 2014; 9:423-436. [PMID: 25580106 DOI: 10.1007/s11515-014-1332-0] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022]
Abstract
Between 15-20% of human cancers are associated with infection by oncogenic viruses. Oncogenic viruses, including HPV, HBV, HCV and HTLV-1, target mitochondria to influence cell proliferation and survival. Oncogenic viral gene products also trigger the production of reactive oxygen species which can elicit oxidative DNA damage and potentiate oncogenic host signaling pathways. Viral oncogenes may also subvert mitochondria quality control mechanisms such as mitophagy and metabolic adaptation pathways to promote virus replication. Here, we will review recent progress on viral regulation of mitophagy and metabolic adaptation and their roles in viral oncogenesis.
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Affiliation(s)
- Young Bong Choi
- Department of Oncology, Sidney Kimmel Comprehensive Cancer Center, Johns HopkinsSchool of Medicine, Baltimore, MD 21287, USA
| | - Edward William Harhaj
- Department of Oncology, Sidney Kimmel Comprehensive Cancer Center, Johns HopkinsSchool of Medicine, Baltimore, MD 21287, USA
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35
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Eggert D, Rösch K, Reimer R, Herker E. Visualization and analysis of hepatitis C virus structural proteins at lipid droplets by super-resolution microscopy. PLoS One 2014; 9:e102511. [PMID: 25019511 PMCID: PMC4094509 DOI: 10.1371/journal.pone.0102511] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/06/2014] [Accepted: 06/19/2014] [Indexed: 12/17/2022] Open
Abstract
Cytosolic lipid droplets are central organelles in the Hepatitis C Virus (HCV) life cycle. The viral capsid protein core localizes to lipid droplets and initiates the production of viral particles at lipid droplet–associated ER membranes. Core is thought to encapsidate newly synthesized viral RNA and, through interaction with the two envelope proteins E1 and E2, bud into the ER lumen. Here, we visualized the spatial distribution of HCV structural proteins core and E2 in vicinity of small lipid droplets by three-color 3D super-resolution microscopy. We observed and analyzed small areas of colocalization between the two structural proteins in HCV-infected cells with a diameter of approximately 100 nm that might represent putative viral assembly sites.
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Affiliation(s)
- Dennis Eggert
- Heinrich-Pette-Institute, Leibniz Institute for Experimental Virology, University of Hamburg, Hamburg, Germany
- Institute of Physical Chemistry, University of Hamburg, Hamburg, Germany
| | - Kathrin Rösch
- Heinrich-Pette-Institute, Leibniz Institute for Experimental Virology, University of Hamburg, Hamburg, Germany
| | - Rudolph Reimer
- Heinrich-Pette-Institute, Leibniz Institute for Experimental Virology, University of Hamburg, Hamburg, Germany
| | - Eva Herker
- Heinrich-Pette-Institute, Leibniz Institute for Experimental Virology, University of Hamburg, Hamburg, Germany
- * E-mail:
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36
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Kalinina OV. GENOME ORGANIZATION AND GEOGRAPHICAL DISTRIBUTION OF THE NATURAL INTERGENOTYPIC RECOMBINANT OF HEPATITIS C VIRUS RF1_2k/1b. ACTA ACUST UNITED AC 2014. [DOI: 10.15789/2220-7619-2012-4-677-686] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/01/2022]
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Abstract
The lifecycle of several viruses is intimately tied to the lipid metabolism of their host cells, and lipid droplets (LDs) have emerged as crucial organelles in the propagation of these viral infections. Investigating the roles of LDs in viral infection requires expertise in both virology and cell metabolism pertaining to LDs. In this review, we offer an updated list and review of the multiples methods we have used in our laboratory to study both the role of LDs in viral infection and the effect of viral infection on cellular LDs, with a special emphasis on hepatitis C virus and other RNA viruses.
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Hino K, Hara Y, Nishina S. Mitochondrial reactive oxygen species as a mystery voice in hepatitis C. Hepatol Res 2014; 44:123-32. [PMID: 24112394 DOI: 10.1111/hepr.12247] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/25/2013] [Revised: 09/05/2013] [Accepted: 09/19/2013] [Indexed: 12/21/2022]
Abstract
There are several lines of evidence suggesting that oxidative stress is present in hepatitis C to a greater degree than in other inflammatory liver diseases and is closely related to disease progression. The main production site of reactive oxygen species (ROS) is assumed to be mitochondria, which concept is supported by evidence that hepatitis C virus (HCV) core protein is directly associated with them. The detoxification of ROS also is an important function of the cellular redox homeostasis system. These results draw our attention to how HCV-induced mitochondrial ROS production is beyond redox regulation and affects the disease progression and development of hepatocellular carcinoma (HCC) in chronic hepatitis C. On the other hand, HCV-related chronic liver diseases are characterized by metabolic alterations such as insulin resistance, hepatic steatosis and/or iron accumulation in the liver. These metabolic disorders also are relevant to the development of HCC in HCV-related chronic liver diseases. Here, we review the mechanisms by which HCV increases mitochondrial ROS production and offer new insights as to how mitochondrial ROS are linked to metabolic disorders such as insulin resistance, hepatic steatosis and hepatic iron accumulation that are observed in HCV-related chronic liver diseases.
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Affiliation(s)
- Keisuke Hino
- Department of Hepatology and Pancreatology, Kawasaki Medical School, Kurashiki, Japan
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Arciello M, Gori M, Balsano C. Mitochondrial dysfunctions and altered metals homeostasis: new weapons to counteract HCV-related oxidative stress. OXIDATIVE MEDICINE AND CELLULAR LONGEVITY 2013; 2013:971024. [PMID: 24371505 PMCID: PMC3859171 DOI: 10.1155/2013/971024] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 06/10/2013] [Revised: 10/18/2013] [Accepted: 10/28/2013] [Indexed: 02/06/2023]
Abstract
The hepatitis C virus (HCV) infection produces several pathological effects in host organism through a wide number of molecular/metabolic pathways. Today it is worldwide accepted that oxidative stress actively participates in HCV pathology, even if the antioxidant therapies adopted until now were scarcely effective. HCV causes oxidative stress by a variety of processes, such as activation of prooxidant enzymes, weakening of antioxidant defenses, organelle damage, and metals unbalance. A focal point, in HCV-related oxidative stress onset, is the mitochondrial failure. These organelles, known to be the "power plants" of cells, have a central role in energy production, metabolism, and metals homeostasis, mainly copper and iron. Furthermore, mitochondria are direct viral targets, because many HCV proteins associate with them. They are the main intracellular free radicals producers and targets. Mitochondrial dysfunctions play a key role in the metal imbalance. This event, today overlooked, is involved in oxidative stress exacerbation and may play a role in HCV life cycle. In this review, we summarize the role of mitochondria and metals in HCV-related oxidative stress, highlighting the need to consider their deregulation in the HCV-related liver damage and in the antiviral management of patients.
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Affiliation(s)
- Mario Arciello
- Department of Internal Medicine and Medical Specialties, “Sapienza” University of Rome, Via del Policlinico 155, 00161 Rome, Italy
- Francesco Balsano Foundation, Via G.B. Martini 6, 00198 Rome, Italy
| | - Manuele Gori
- Francesco Balsano Foundation, Via G.B. Martini 6, 00198 Rome, Italy
| | - Clara Balsano
- Francesco Balsano Foundation, Via G.B. Martini 6, 00198 Rome, Italy
- Institute of Molecular Biology and Pathology (IBPM); CNR, Piazzale Aldo Moro 7, 00185 Rome, Italy
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Neufeldt CJ, Joyce MA, Levin A, Steenbergen RH, Pang D, Shields J, Tyrrell DLJ, Wozniak RW. Hepatitis C virus-induced cytoplasmic organelles use the nuclear transport machinery to establish an environment conducive to virus replication. PLoS Pathog 2013; 9:e1003744. [PMID: 24204278 PMCID: PMC3814334 DOI: 10.1371/journal.ppat.1003744] [Citation(s) in RCA: 50] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/14/2012] [Accepted: 09/19/2013] [Indexed: 12/12/2022] Open
Abstract
Hepatitis C virus (HCV) infection induces formation of a membranous web structure in the host cell cytoplasm where the viral genome replicates and virions assemble. The membranous web is thought to concentrate viral components and hide viral RNA from pattern recognition receptors. We have uncovered a role for nuclear pore complex proteins (Nups) and nuclear transport factors (NTFs) in the membranous web. We show that HCV infection leads to increased levels of cytoplasmic Nups that accumulate at sites enriched for HCV proteins. Moreover, we detected interactions between specific HCV proteins and both Nups and NTFs. We hypothesize that cytoplasmically positioned Nups facilitate formation of the membranous web and contribute to the compartmentalization of viral replication. Accordingly, we show that transport cargo proteins normally targeted to the nucleus are capable of entering regions of the membranous web, and that depletion of specific Nups or Kaps inhibits HCV replication and assembly.
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Affiliation(s)
| | - Michael A. Joyce
- Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada
| | - Aviad Levin
- Department of Cell Biology, University of Alberta, Edmonton, Alberta, Canada
- Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada
| | - Rineke H. Steenbergen
- Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada
| | - Daniel Pang
- Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada
| | - Justin Shields
- Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada
| | - D. Lorne J. Tyrrell
- Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada
| | - Richard W. Wozniak
- Department of Cell Biology, University of Alberta, Edmonton, Alberta, Canada
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41
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Blum HE. Hepatitis C and Hepatocellular Carcinoma. VIRAL HEPATITIS 2013:353-361. [DOI: 10.1002/9781118637272.ch24] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/04/2025]
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Wang T, Weinman SA. Interactions Between Hepatitis C Virus and Mitochondria: Impact on Pathogenesis and Innate Immunity. CURRENT PATHOBIOLOGY REPORTS 2013; 1:179-187. [PMID: 23956955 DOI: 10.1007/s40139-013-0024-9] [Citation(s) in RCA: 31] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Hepatitis C virus (HCV) causes a persistent chronic infection of hepatocytes resulting in progressive fibrosis and carcinogenesis. Abnormalities in mitochondria are prominent features of clinical disease where ultrastructural changes, alterations in electron transport, and excess reactive oxygen species (ROS) production occur. These mitochondrial abnormalities correlate with disease severity and resolve with viral eradication. Multiple viral proteins, particularly core and NS3/4a bind to mitochondria. The core and NS5a proteins primarily cause ER stress, ER Ca2+ release and enhance direct transfer of Ca2+ from ER to mitochondria. This results in electron transport changes, increased ROS production and sensitivity to mitochondrial permeability transition and cell death. The viral protease, NS3/4a, binds to mitochondria as well where it cleaves an important signaling adapter, MAVS, thus preventing viral clearance by endogenous interferon production. This review discusses the mechanisms by which HCV causes mitochondrial changes and consequences of these for disease.
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Affiliation(s)
- Ting Wang
- Liver Center and Department of Internal Medicine, University of Kansas Medical Center, Kansas City, KS 66160
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43
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Mazumder N, Lyn RK, Singaravelu R, Ridsdale A, Moffatt DJ, Hu CW, Tsai HR, McLauchlan J, Stolow A, Kao FJ, Pezacki JP. Fluorescence lifetime imaging of alterations to cellular metabolism by domain 2 of the hepatitis C virus core protein. PLoS One 2013; 8:e66738. [PMID: 23826122 PMCID: PMC3691201 DOI: 10.1371/journal.pone.0066738] [Citation(s) in RCA: 25] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/07/2013] [Accepted: 05/09/2013] [Indexed: 12/15/2022] Open
Abstract
Hepatitis C virus (HCV) co-opts hepatic lipid pathways to facilitate its pathogenesis. The virus alters cellular lipid biosynthesis and trafficking, and causes an accumulation of lipid droplets (LDs) that gives rise to hepatic steatosis. Little is known about how these changes are controlled at the molecular level, and how they are related to the underlying metabolic states of the infected cell. The HCV core protein has previously been shown to independently induce alterations in hepatic lipid homeostasis. Herein, we demonstrate, using coherent anti-Stokes Raman scattering (CARS) microscopy, that expression of domain 2 of the HCV core protein (D2) fused to GFP is sufficient to induce an accumulation of larger lipid droplets (LDs) in the perinuclear region. Additionally, we performed fluorescence lifetime imaging of endogenous reduced nicotinamide adenine dinucleotides [NAD(P)H], a key coenzyme in cellular metabolic processes, to monitor changes in the cofactor’s abundance and conformational state in D2-GFP transfected cells. When expressed in Huh-7 human hepatoma cells, we observed that the D2-GFP induced accumulation of LDs correlated with an increase in total NAD(P)H fluorescence and an increase in the ratio of free to bound NAD(P)H. This is consistent with an approximate 10 fold increase in cellular NAD(P)H levels. Furthermore, the lifetimes of bound and free NAD(P)H were both significantly reduced – indicating viral protein-induced alterations in the cofactors’ binding and microenvironment. Interestingly, the D2-expressing cells showed a more diffuse localization of NAD(P)H fluorescence signal, consistent with an accumulation of the co-factor outside the mitochondria. These observations suggest that HCV causes a shift of metabolic control away from the use of the coenzyme in mitochondrial electron transport and towards glycolysis, lipid biosynthesis, and building of new biomass. Overall, our findings demonstrate that HCV induced alterations in hepatic metabolism is tightly linked to alterations in NAD(P)H functional states.
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Affiliation(s)
- Nirmal Mazumder
- Institute of Biophotonics, National Yang-Ming University, Taipei, Taiwan
| | - Rodney K. Lyn
- National Research Council of Canada, Ottawa, Ontario, Canada
- Department of Chemistry, University of Ottawa, Ottawa, Ontario, Canada
| | - Ragunath Singaravelu
- National Research Council of Canada, Ottawa, Ontario, Canada
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, Ontario, Canada
| | - Andrew Ridsdale
- National Research Council of Canada, Ottawa, Ontario, Canada
| | | | - Chih-Wei Hu
- Institute of Biophotonics, National Yang-Ming University, Taipei, Taiwan
| | - Han-Ruei Tsai
- Institute of Biophotonics, National Yang-Ming University, Taipei, Taiwan
| | - John McLauchlan
- Medical Research Council - University of Glasgow Center for Virus Research, Glasgow, United Kingdom
| | - Albert Stolow
- National Research Council of Canada, Ottawa, Ontario, Canada
- Department of Physics, Queen’s University, Kingston, Ontario, Canada
| | - Fu-Jen Kao
- Institute of Biophotonics, National Yang-Ming University, Taipei, Taiwan
- * E-mail: (JPP); (FK)
| | - John Paul Pezacki
- National Research Council of Canada, Ottawa, Ontario, Canada
- Department of Chemistry, University of Ottawa, Ottawa, Ontario, Canada
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, Ontario, Canada
- * E-mail: (JPP); (FK)
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Hepatitis C virus induces the mitochondrial translocation of Parkin and subsequent mitophagy. PLoS Pathog 2013; 9:e1003285. [PMID: 23555273 PMCID: PMC3610669 DOI: 10.1371/journal.ppat.1003285] [Citation(s) in RCA: 150] [Impact Index Per Article: 12.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/19/2012] [Accepted: 02/19/2013] [Indexed: 12/16/2022] Open
Abstract
Hepatitis C Virus (HCV) induces intracellular events that trigger mitochondrial dysfunction and promote host metabolic alterations. Here, we investigated selective autophagic degradation of mitochondria (mitophagy) in HCV-infected cells. HCV infection stimulated Parkin and PINK1 gene expression, induced perinuclear clustering of mitochondria, and promoted mitochondrial translocation of Parkin, an initial event in mitophagy. Liver tissues from chronic HCV patients also exhibited notable levels of Parkin induction. Using multiple strategies involving confocal and electron microscopy, we demonstrated that HCV-infected cells display greater number of mitophagosomes and mitophagolysosomes compared to uninfected cells. HCV-induced mitophagy was evidenced by the colocalization of LC3 puncta with Parkin-associated mitochondria and lysosomes. Ultrastructural analysis by electron microscopy and immunoelectron microscopy also displayed engulfment of damaged mitochondria in double membrane vesicles in HCV-infected cells. The HCV-induced mitophagy occurred irrespective of genotypic differences. Silencing Parkin and PINK1 hindered HCV replication suggesting the functional relevance of mitophagy in HCV propagation. HCV-mediated decline of mitochondrial complex I enzyme activity was rescued by chemical inhibition of mitophagy or by Parkin silencing. Overall our results suggest that HCV induces Parkin-dependent mitophagy, which may have significant contribution in mitochondrial liver injury associated with chronic hepatitis C. Hepatitis C virus (HCV) infection alters host lipid metabolism. HCV-induced mitochondrial dysfunction may promote the metabolic alterations by affecting mitochondrial β-oxidation and oxidative phosphorylation. Dysfunctional mitochondria are detrimental to cell survival and require rapid clearance to sustain cell viability. Here, we investigated the effect of HCV gene expression in promoting selective autophagy of dysfunctional mitochondria, also termed mitophagy. HCV infection stimulated the gene expression of Parkin and PINK1, the two key mediators of mitophagy. Parkin stimulation was also observed in liver biopsies of chronic hepatitis C patients. HCV infection induced the perinuclear clustering of mitochondria and triggered Parkin translocation to mitochondria, a hallmark of mitophagy. Concomitant with the mitochondrial translocation of Parkin, we observed ubiquitination of Parkin and its substrates in HCV-infected cells. We also demonstrate the formation of mitophagosomes and their subsequent delivery to lysosomes in HCV-infected cells. Silencing both Parkin and PINK1 hindered HCV replication, suggesting the functional significance of mitophagy in HCV life cycle. Furthermore, we demonstrate that Parkin-dependent mitophagy is directly associated with HCV-mediated decline in oxidative phosphorylation. Our results implicate the functional significance of Parkin and mitophagy in the persistence of HCV infection and mitochondrial injury commonly seen in patients with chronic hepatitis C.
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Hepatitis C virus-induced mitochondrial dysfunctions. Viruses 2013; 5:954-80. [PMID: 23518579 PMCID: PMC3705306 DOI: 10.3390/v5030954] [Citation(s) in RCA: 50] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/22/2013] [Revised: 03/15/2013] [Accepted: 03/20/2013] [Indexed: 12/15/2022] Open
Abstract
Chronic hepatitis C is characterized by metabolic disorders and a microenvironment in the liver dominated by oxidative stress, inflammation and regeneration processes that lead in the long term to hepatocellular carcinoma. Many lines of evidence suggest that mitochondrial dysfunctions, including modification of metabolic fluxes, generation and elimination of oxidative stress, Ca2+ signaling and apoptosis, play a central role in these processes. However, how these dysfunctions are induced by the virus and whether they play a role in disease progression and neoplastic transformation remains to be determined. Most in vitro studies performed so far have shown that several of the hepatitis C virus (HCV) proteins localize to mitochondria, but the consequences of these interactions on mitochondrial functions remain contradictory, probably due to the use of artificial expression and replication systems. In vivo studies are hampered by the fact that innate and adaptive immune responses will overlay mitochondrial dysfunctions induced directly in the hepatocyte by HCV. Thus, the molecular aspects underlying HCV-induced mitochondrial dysfunctions and their roles in viral replication and the associated pathology need yet to be confirmed in the context of productively replicating virus and physiologically relevant in vitro and in vivo model systems.
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Ivanov AV, Bartosch B, Smirnova OA, Isaguliants MG, Kochetkov SN. HCV and oxidative stress in the liver. Viruses 2013; 5:439-69. [PMID: 23358390 PMCID: PMC3640510 DOI: 10.3390/v5020439] [Citation(s) in RCA: 157] [Impact Index Per Article: 13.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/26/2012] [Revised: 12/26/2012] [Accepted: 01/17/2013] [Indexed: 12/12/2022] Open
Abstract
Hepatitis C virus (HCV) is the etiological agent accounting for chronic liver disease in approximately 2-3% of the population worldwide. HCV infection often leads to liver fibrosis and cirrhosis, various metabolic alterations including steatosis, insulin and interferon resistance or iron overload, and development of hepatocellular carcinoma or non-Hodgkin lymphoma. Multiple molecular mechanisms that trigger the emergence and development of each of these pathogenic processes have been identified so far. One of these involves marked induction of a reactive oxygen species (ROS) in infected cells leading to oxidative stress. To date, markers of oxidative stress were observed both in chronic hepatitis C patients and in various in vitro systems, including replicons or stable cell lines expressing viral proteins. The search for ROS sources in HCV-infected cells revealed several mechanisms of ROS production and thus a number of cellular proteins have become targets for future studies. Furthermore, during last several years it has been shown that HCV modifies antioxidant defense mechanisms. The aim of this review is to summarize the present state of art in the field and to try to predict directions for future studies.
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Affiliation(s)
- Alexander V. Ivanov
- Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilov str., 32, Moscow 119991, Russia; E-Mails: (A.I.); (O.S.); (S.K.)
| | - Birke Bartosch
- CRCL, INSERM U1052, CNRS 5286, Université de Lyon, 151, Cours A Thomas 69424 Lyon Cedex France; E-Mail:
| | - Olga A. Smirnova
- Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilov str., 32, Moscow 119991, Russia; E-Mails: (A.I.); (O.S.); (S.K.)
| | - Maria G. Isaguliants
- Department of Molecular Biology, Tumor and Cell Biology, Karolinska Institutet, Nobels väg 16 17177 Stockholm, Sweden; E-Mail:
- D.I. Ivanovsky Institute of Virology, Gamaleya Str. 16, 123098 Moscow, Russia; E-Mail:
| | - Sergey N. Kochetkov
- Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilov str., 32, Moscow 119991, Russia; E-Mails: (A.I.); (O.S.); (S.K.)
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Terasaki K, Won S, Makino S. The C-terminal region of Rift Valley fever virus NSm protein targets the protein to the mitochondrial outer membrane and exerts antiapoptotic function. J Virol 2013; 87:676-82. [PMID: 23097454 PMCID: PMC3536385 DOI: 10.1128/jvi.02192-12] [Citation(s) in RCA: 42] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/16/2012] [Accepted: 10/18/2012] [Indexed: 11/20/2022] Open
Abstract
The NSm nonstructural protein of Rift Valley fever virus (family Bunyaviridae, genus Phlebovirus) has an antiapoptotic function and affects viral pathogenesis. We found that NSm integrates into the mitochondrial outer membrane and that the protein's N terminus is exposed to the cytoplasm. The C-terminal region of NSm, which contains a basic amino acid cluster and a putative transmembrane domain, targeted the protein to the mitochondrial outer membrane and exerted antiapoptotic function.
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Affiliation(s)
| | | | - Shinji Makino
- Department of Microbiology and Immunology
- Center for Biodefense and Emerging Infectious Diseases
- UTMB Center for Tropical Diseases
- Sealy Center for Vaccine Development, The University of Texas Medical Branch, Galveston, Texas, USA
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48
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PPARs and HCV-Related Hepatocarcinoma: A Mitochondrial Point of View. PPAR Res 2012; 2012:605302. [PMID: 22966221 PMCID: PMC3431172 DOI: 10.1155/2012/605302] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/08/2012] [Accepted: 07/16/2012] [Indexed: 12/24/2022] Open
Abstract
Hepatitis-C-virus-related infective diseases are worldwide spread pathologies affecting primarily liver. The infection is often asymptomatic, but when chronically persisting can lead to liver scarring and ultimately to cirrhosis, which is generally apparent after decades. In some cases, cirrhosis will progress to develop liver failure, liver cancer, or life-threatening esophageal and gastric varices. HCV-infected cells undergo profound metabolic dysregulation whose mechanisms are yet not well understood. An emerging feature in the pathogenesis of the HCV-related disease is the setting of a pro-oxidative condition caused by dysfunctions of mitochondria which proved to be targets of viral proteins. This causes deregulation of mitochondria-dependent catabolic pathway including fatty acid oxidation. Nuclear receptors and their ligands are fundamental regulators of the liver metabolic homeostasis, which are disrupted following HCV infection. In this contest, specific attention has been focused on the peroxisome proliferator activated receptors given their role in controlling liver lipid metabolism and the availability of specific pharmacological drugs of potential therapeutic utilization. However, the reported role of PPARs in HCV infection provides conflicting results likely due to different species-specific contests. In this paper we summarize the current knowledge on this issue and offer a reconciling model based on mitochondria-related features.
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Quarato G, Scrima R, Agriesti F, Moradpour D, Capitanio N, Piccoli C. Targeting mitochondria in the infection strategy of the hepatitis C virus. Int J Biochem Cell Biol 2012; 45:156-66. [PMID: 22710347 DOI: 10.1016/j.biocel.2012.06.008] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/13/2012] [Revised: 05/29/2012] [Accepted: 06/07/2012] [Indexed: 12/23/2022]
Abstract
Hepatitis C virus (HCV) infection induces a state of oxidative stress more pronounced than that observed in many other inflammatory diseases. Here, we propose a temporal sequence of events in the HCV-infected cell whereby the primary alteration consists of a release of Ca(2+) from the endoplasmic reticulum, followed by uptake into mitochondria. This ensues successive mitochondrial dysfunction leading to the generation of reactive oxygen species and a progressive metabolic adaptive response. Evidence is provided for a positive feed-back mechanism between alterations of calcium and redox homeostasis. This likely involves deregulation of the mitochondrial permeability transition and induces progressive dysfunction of cellular bioenergetics. Pathogenetic implications of the model and new opportunities for therapeutic intervention are discussed. This article is part of a Directed Issue entitled: Bioenergetic dysfunction, adaptation and therapy.
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Affiliation(s)
- Giovanni Quarato
- Department of Biomedical Sciences, University of Foggia, Foggia, Italy
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50
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HIV-1 Vpr triggers mitochondrial destruction by impairing Mfn2-mediated ER-mitochondria interaction. PLoS One 2012; 7:e33657. [PMID: 22438978 PMCID: PMC3306277 DOI: 10.1371/journal.pone.0033657] [Citation(s) in RCA: 70] [Impact Index Per Article: 5.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2011] [Accepted: 02/17/2012] [Indexed: 11/19/2022] Open
Abstract
Human immunodeficiency virus 1 (HIV-1) viral protein R (Vpr) has been shown to induce host cell death by increasing the permeability of mitochondrial outer membrane (MOM). The mechanism underlying the damage to the mitochondria by Vpr, however, is not clearly illustrated. In this study, Vpr that is introduced, via transient transfection or lentivirus infection, into the human embryonic kidney cell line HEK293, human CD4+ T lymphoblast cell line SupT1, or human primary CD4+ T cells serves as the model system to study the molecular mechanism of Vpr-mediated HIV-1 pathogenesis. The results show that Vpr injures MOM and causes a loss in membrane potential (MMP) by posttranscriptionally reducing the expression of mitofusin 2 (Mfn2) via VprBP-DDB1-CUL4A ubiquitin ligase complex, gradually weakening MOM, and increasing mitochondrial deformation. Vpr also markedly decreases cytoplasmic levels of dynamin-related protein 1 (DRP1) and increases bulging in mitochondria-associated membranes (MAM), the specific regions of endoplasmic reticulum (ER) which form physical contacts with the mitochondria. Overexpression of Mfn2 and DRP1 significantly decreased the loss of MMP and apoptotic cell death caused by Vpr. Furthermore, by employing time-lapse confocal fluorescence microscopy, we identify the transport of Vpr protein from the ER, via MAM to the mitochondria. Taken together, our results suggest that Vpr-mediated cellular damage may occur on an alternative protein transport pathway from the ER, via MAM to the mitochondria, which are modulated by Mfn2 and DRP1.
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