scholarly journals Ectopic Expression of Hepatitis C Virus Core Protein Differentially Regulates Nuclear Transcription Factors

1998 ◽  
Vol 72 (12) ◽  
pp. 9722-9728 ◽  
Author(s):  
Anju Shrivastava ◽  
Sunil K. Manna ◽  
Ranjit Ray ◽  
Bharat B. Aggarwal
Keyword(s):  
Transcription Factors ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Inflammatory Responses ◽  
Core Protein ◽  
Ectopic Expression ◽  
Cellular Growth ◽  
Mitogen Activated Protein Kinase ◽  
The Core ◽  
Hcv Core Protein

ABSTRACT The putative core protein of hepatitis C virus (HCV) regulates cellular growth and a number of cellular promoters. To further understand its effect, we investigated the role of the core protein in the endogenous regulation of two distinct transcription factors, nuclear factor-κB (NF-κB) and activating protein-1 (AP-1), and the related mitogen-activated protein kinase kinase (MAPKK) and c-Jun N-terminal kinase (JNK). Stable cell transfectants expressing the HCV core protein suppressed tumor necrosis factor (TNF)-induced NF-κB activation. Supershift analysis revealed that NF-κB consists of p50 and p65 subunits. This correlated with inhibition of the degradation of IκBα, the inhibitory subunit of NF-κB. The effect was not specific to TNF, as suppression in core protein-expressing cells was also observed in response to a number of other inflammatory agents known to activate NF-κB. In contrast to the effect on NF-κB, the HCV core protein constitutively activated AP-1, which correlated with the activation of JNK and MAPKK, which are known to regulate AP-1. These observations indicated that the core protein targets transcription factors known to be involved in the regulation of inflammatory responses and the immune system.

scholarly journals Interaction of Hepatitis C Virus Core Protein with Viral Sense RNA and Suppression of Its Translation

1999 ◽  
Vol 73 (12) ◽  
pp. 9718-9725 ◽  
Author(s):  
Takashi Shimoike ◽  
Shigetaka Mimori ◽  
Hideki Tani ◽  
Yoshiharu Matsuura ◽  
Tatsuo Miyamura
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Core Protein ◽  
Firefly Luciferase ◽  
Encephalomyocarditis Virus ◽  
Hcv Rna ◽  
Dependent Manner ◽  
Genomic Rna ◽  
The Core ◽  
Hcv Core Protein

ABSTRACT To clarify the binding properties of hepatitis C virus (HCV) core protein and its viral RNA for the encapsidation, morphogenesis, and replication of HCV, the specific interaction of HCV core protein with its genomic RNA synthesized in vitro was examined in an in vivo system. The positive-sense RNA from the 5′ end to nucleotide (nt) 2327, which covers the 5′ untranslated region (5′UTR) and a part of the coding region of HCV structural proteins, interacted with HCV core protein, while no interaction was observed in the same region of negative-sense RNA and in other regions of viral and antiviral sense RNAs. The internal ribosome entry site (IRES) exists around the 5′UTR of HCV; therefore, the interaction of the core protein with this region of HCV RNA suggests that there is some effect on its cap-independent translation. Cells expressing HCV core protein were transfected with reporter RNAs consisting of nt 1 to 709 of HCV RNA (the 5′UTR of HCV and about two-thirds of the core protein coding regions) followed by a firefly luciferase gene (HCV07Luc RNA). The translation of HCV07Luc RNA was suppressed in cells expressing the core protein, whereas no significant suppression was observed in the case of a reporter RNA possessing the IRES of encephalomyocarditis virus followed by a firefly luciferase. This suppression by the core protein occurred in a dose-dependent manner. The expression of the E1 envelope protein of HCV or β-galactosidase did not suppress the translation of both HCV and EMCV reporter RNAs. We then examined the regions that are important for suppression of translation by the core protein and found that the region from nt 1 to 344 was enough to exert this suppression. These results suggest that the HCV core protein interacts with viral genomic RNA at a specific region to form nucleocapsids and regulates the expression of HCV by interacting with the 5′UTR.

scholarly journals Intramembrane Processing by Signal Peptide Peptidase Regulates the Membrane Localization of Hepatitis C Virus Core Protein and Viral Propagation

2008 ◽  
Vol 82 (17) ◽  
pp. 8349-8361 ◽  
Author(s):  
Kiyoko Okamoto ◽  
Yoshio Mori ◽  
Yasumasa Komoda ◽  
Toru Okamoto ◽  
Masayasu Okochi ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Signal Peptide ◽  
Core Protein ◽  
Biological Significance ◽  
Signal Peptidase ◽  
Signal Peptide Peptidase ◽  
Viral Propagation ◽  
The Core ◽  
Hcv Core Protein

ABSTRACT Hepatitis C virus (HCV) core protein has shown to be localized in the detergent-resistant membrane (DRM), which is distinct from the classical raft fraction including caveolin, although the biological significance of the DRM localization of the core protein has not been determined. The HCV core protein is cleaved off from a precursor polyprotein at the lumen side of Ala191 by signal peptidase and is then further processed by signal peptide peptidase (SPP) within the transmembrane region. In this study, we examined the role of SPP in the localization of the HCV core protein in the DRM and in viral propagation. The C terminus of the HCV core protein cleaved by SPP in 293T cells was identified as Phe177 by mass spectrometry. Mutations introduced into two residues (Ile176 and Phe177) upstream of the cleavage site of the core protein abrogated processing by SPP and localization in the DRM fraction. Expression of a dominant-negative SPP or treatment with an SPP inhibitor, L685,458, resulted in reductions in the levels of processed core protein localized in the DRM fraction. The production of HCV RNA in cells persistently infected with strain JFH-1 was impaired by treatment with the SPP inhibitor. Furthermore, mutant JFH-1 viruses bearing SPP-resistant mutations in the core protein failed to propagate in a permissive cell line. These results suggest that intramembrane processing of HCV core protein by SPP is required for the localization of the HCV core protein in the DRM and for viral propagation.

scholarly journals Hepatitis C Virus Genotype 1b Core Protein Does Not Exert Immunomodulatory Effects on Virus-Induced Cellular Immunity

2002 ◽  
Vol 76 (3) ◽  
pp. 990-997 ◽  
Author(s):  
Zhang-Xu Liu ◽  
Hiroshi Nishida ◽  
Jian-Wen He ◽  
Michael M. C. Lai ◽  
Ni Feng ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Liver Injury ◽  
Core Protein ◽  
Virus Genotype ◽  
Hcv Genotype ◽  
Cell Functions ◽  
The Core ◽  
Genotype 1B ◽  
Hcv Core Protein

ABSTRACT The hepatitis C virus (HCV) core protein is among the most conserved proteins in HCV and is known to induce sensitization of cytotoxic T lymphocytes (CTL). Therefore, it is a prime candidate for a component of a potential HCV vaccine. The HCV core protein has, however, been reported to exert multiple effects on cell functions, raising questions as to its suitability for this purpose. This question was investigated here with mice into which replication-deficient adenoviruses expressing core protein of an HCV genotype 1b isolate were injected. We show that induction of cytokines in response to the infection, infiltration of lymphocytes into the infected liver, priming of virus-specific CTL, and liver injury are not modulated by expression of the core protein in the liver. Moreover, no changes in the sensitivity to tumor necrosis factor alpha- or Fas-mediated liver injury are demonstrable. A similar lack of demonstrable effects of the core protein on immune functions has also been obtained using transgenic mice expressing another HCV genotype 1b core protein. It is concluded that the HCV core protein of genotype 1b has no modulatory effects on induction of virus-specific immune responses and may therefore be a suitable component of an HCV vaccine.

scholarly journals Characterization of Hepatitis C Virus Core Protein Multimerization and Membrane Envelopment: Revelation of a Cascade of Core-Membrane Interactions

2009 ◽  
Vol 83 (19) ◽  
pp. 9923-9939 ◽  
Author(s):  
Li-Shuang Ai ◽  
Yu-Wen Lee ◽  
Steve S.-L. Chen
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Complex Formation ◽  
Signal Peptide ◽  
Core Protein ◽  
The Core ◽  
Hcv Core Protein ◽  
Core Proteins ◽  
Multimeric Complex ◽  
Protein Multimerization

ABSTRACT The molecular basis underlying hepatitis C virus (HCV) core protein maturation and morphogenesis remains elusive. We characterized the concerted events associated with core protein multimerization and interaction with membranes. Analyses of core proteins expressed from a subgenomic system showed that the signal sequence located between the core and envelope glycoprotein E1 is critical for core association with endoplasmic reticula (ER)/late endosomes and the core's envelopment by membranes, which was judged by the core's acquisition of resistance to proteinase K digestion. Despite exerting an inhibitory effect on the core's association with membranes, (Z-LL)2-ketone, a specific inhibitor of signal peptide peptidase (SPP), did not affect core multimeric complex formation, suggesting that oligomeric core complex formation proceeds prior to or upon core attachment to membranes. Protease-resistant core complexes that contained both innate and processed proteins were detected in the presence of (Z-LL)2-ketone, implying that core envelopment occurs after intramembrane cleavage. Mutations of the core that prevent signal peptide cleavage or coexpression with an SPP loss-of-function D219A mutant decreased the core's envelopment, demonstrating that SPP-mediated cleavage is required for core envelopment. Analyses of core mutants with a deletion in domain I revealed that this domain contains sequences crucial for core envelopment. The core proteins expressed by infectious JFH1 and Jc1 RNAs in Huh7 cells also assembled into a multimeric complex, associated with ER/late-endosomal membranes, and were enveloped by membranes. Treatment with (Z-LL)2-ketone or coexpression with D219A mutant SPP interfered with both core envelopment and infectious HCV production, indicating a critical role of core envelopment in HCV morphogenesis. The results provide mechanistic insights into the sequential and coordinated processes during the association of the HCV core protein with membranes in the early phase of virus maturation and morphogenesis.

scholarly journals Molecular Determinants for Subcellular Localization of Hepatitis C Virus Core Protein

2005 ◽  
Vol 79 (2) ◽  
pp. 1271-1281 ◽  
Author(s):  
Ryosuke Suzuki ◽  
Shinichiro Sakamoto ◽  
Takeya Tsutsumi ◽  
Akiko Rikimaru ◽  
Keiko Tanaka ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Nuclear Translocation ◽  
Core Protein ◽  
Mitochondrial Outer Membrane ◽  
Cellular Trafficking ◽  
Nuclear Targeting ◽  
Diverse Range ◽  
The Core ◽  
Hcv Core Protein

ABSTRACT Hepatitis C virus (HCV) core protein is a putative nucleocapsid protein with a number of regulatory functions. In tissue culture cells, HCV core protein is mainly located at the endoplasmic reticulum as well as mitochondria and lipid droplets within the cytoplasm. However, it is also detected in the nucleus in some cells. To elucidate the mechanisms by which cellular trafficking of the protein is controlled, we performed subcellular fractionation experiments and used confocal microscopy to examine the distribution of heterologously expressed fusion proteins involving various deletions and point mutations of the HCV core combined with green fluorescent proteins. We demonstrated that a region spanning amino acids 112 to 152 can mediate association of the core protein not only with the ER but also with the mitochondrial outer membrane. This region contains an 18-amino-acid motif which is predicted to form an amphipathic α-helix structure. With regard to the nuclear targeting of the core protein, we identified a novel bipartite nuclear localization signal, which requires two out of three basic-residue clusters for efficient nuclear translocation, possibly by occupying binding sites on importin-α. Differences in the cellular trafficking of HCV core protein, achieved and maintained by multiple targeting functions as mentioned above, may in part regulate the diverse range of biological roles of the core protein.

scholarly journals Hepatitis C Virus Differentially Modulates Activation of Forkhead Transcription Factors and Insulin-Induced Metabolic Gene Expression

2010 ◽  
Vol 84 (12) ◽  
pp. 5936-5946 ◽  
Author(s):  
Arup Banerjee ◽  
Keith Meyer ◽  
Budhaditya Mazumdar ◽  
Ratna B. Ray ◽  
Ranjit Ray
Keyword(s):  
Gene Expression ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Core Protein ◽  
Ectopic Expression ◽  
Human Hepatocytes ◽  
Hcv Infection ◽  
Metabolic Gene ◽  
Downstream Target ◽  
The Core

ABSTRACT Chronic hepatitis C virus (HCV) infection is often associated with insulin resistance and hepatic steatosis. Insulin regulates gene expression of key enzymes in glucose and lipid metabolism by modulating the activity of specific Forkhead box transcriptional regulators (FoxO1 and FoxA2) via the phosphatidylinositol 3-kinase (PI3K)-Akt signaling pathway in the liver. In this study, we observed that HCV infection of human hepatocytes impaired insulin-induced FoxO1 translocation from the nucleus to the cytoplasm and significantly reduced accumulation of FoxA2 in the nucleus. Phosphorylation of FoxO1 at Ser256, a downstream target for Akt, was inhibited in hepatocytes infected with HCV or expressing the core protein or full-length (FL) genome of HCV. Further, an interaction between FoxO1 and 14-3-3 protein, important for FoxO1 translocation, was inhibited in HCV core-expressing cells. Hepatocytes infected with HCV, expressing the core protein alone or polyprotein displayed an increased level of glucose-6-phosphatase (G6P) mRNA. On the other hand, microsomal triglycerol transfer protein (MTP) activity and apolipoprotein B (ApoB) secretion were significantly reduced in hepatocytes expressing HCV proteins. Together, these observations suggest that HCV infection or ectopic expression of the core protein either alone or together with other viral proteins from an FL gene construct differentially modulates FoxO1 and FoxA2 activation and affects insulin-induced metabolic gene regulation in human hepatocytes.

scholarly journals Hepatitis C Virus Core from Two Different Genotypes Has an Oncogenic Potential but Is Not Sufficient for Transforming Primary Rat Embryo Fibroblasts in Cooperation with the H-ras Oncogene

1998 ◽  
Vol 72 (4) ◽  
pp. 3060-3065 ◽  
Author(s):  
Jun Chang ◽  
Se-Hwan Yang ◽  
Young-Gyu Cho ◽  
Soon Bong Hwang ◽  
Young Shin Hahn ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Cell Line ◽  
Core Protein ◽  
Core Gene ◽  
Primary Cells ◽  
Rat Embryo ◽  
Oncogenic Potential ◽  
The Core ◽  
Hcv Core Protein

ABSTRACT Persistent infection with hepatitis C virus (HCV) is associated with the development of liver cirrhosis and hepatocellular carcinoma. To examine the oncogenic potential of the HCV core gene product, primary rat embryo fibroblasts (REFs) were transfected with the core gene in the presence or absence of the H-ras oncogene. In contrast to a previous report (R. B. Ray, L. M. Lagging, K. Meyer, and R. Ray, J. Virol. 70:4438–4443, 1996), HCV core proteins from two different genotypes (type 1a and type 1b) were not found to transform REFs to tumorigenic phenotype in cooperation with the H-ras oncogene, although the core protein was successfully expressed 20 days after transfection. In addition, REFs transfected with E1A- but not core-expressing plasmid showed the phenotype of immortalized cells when selected with G418. The biological activity was confirmed by observing the transcription activation from two viral promoters, Rous sarcoma virus long terminal repeat and simian virus 40 promoter, which are known to be activated by the core protein from HCV-1 isolate. In contrast to the result with primary cells, the Rat-1 cell line, stably expressing HCV core protein, exhibited focus formation, anchorage-independent growth, and tumor formation in nude mice. HCV core protein was able to induce the transformation of Rat-1 cells with various efficiencies depending on the expression level of the core protein. These results indicate that HCV core protein has an oncogenic potential to transform the Rat-1 cell line but is not sufficient to either immortalize primary REFs by itself or transform primary cells in conjunction with the H-ras oncogene.

scholarly journals Identification of Basic Amino Acids at the N-Terminal End of the Core Protein That Are Crucial for Hepatitis C Virus Infectivity

2010 ◽  
Vol 84 (24) ◽  
pp. 12515-12528 ◽  
Author(s):  
Khaled Alsaleh ◽  
Pierre-Yves Delavalle ◽  
André Pillez ◽  
Gilles Duverlie ◽  
Véronique Descamps ◽  
...  
Keyword(s):  
Amino Acids ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Core Protein ◽  
Basic Amino Acid ◽  
Amino Acid Residues ◽  
The Core ◽  
Hcv Core Protein ◽  
Viral Particles ◽  
A Cell

ABSTRACT A major function of the hepatitis C virus (HCV) core protein is the interaction with genomic RNA to form the nucleocapsid, an essential component of the virus particle. Analyses to identify basic amino acid residues of HCV core protein, important for capsid assembly, were initially performed with a cell-free system, which did not indicate the importance of these residues for HCV infectivity. The development of a cell culture system for HCV (HCVcc) allows a more precise analysis of these core protein amino acids during the HCV life cycle. In the present study, we used a mutational analysis in the context of the HCVcc system to determine the role of the basic amino acid residues of the core protein in HCV infectivity. We focused our analysis on basic residues located in two clusters (cluster 1, amino acids [aa]6 to 23; cluster 2, aa 39 to 62) within the N-terminal 62 amino acids of the HCV core protein. Our data indicate that basic residues of the first cluster have little impact on replication and are dispensable for infectivity. Furthermore, only four basic amino acids residues of the second cluster (R50, K51, R59, and R62) were essential for the production of infectious viral particles. Mutation of these residues did not interfere with core protein subcellular localization, core protein-RNA interaction, or core protein oligomerization. Moreover, these mutations had no effect on core protein envelopment by intracellular membranes. Together, these data indicate that R50, K51, R59, and R62 residues play a major role in the formation of infectious viral particles at a post-nucleocapsid assembly step.

scholarly journals Differential Effects on the Hepatitis C Virus (HCV) Internal Ribosome Entry Site by Vitamin B12 and the HCV Core Protein

2004 ◽  
Vol 78 (21) ◽  
pp. 12075-12081 ◽  
Author(s):  
Dongsheng Li ◽  
William B. Lott ◽  
John Martyn ◽  
Gholamreza Haqshenas ◽  
Eric J. Gowans
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Internal Ribosome Entry Site ◽  
Core Protein ◽  
Vitamin B ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Hcv Core Protein ◽  
Hcv Ires

ABSTRACT To investigate the role of the hepatitis C virus internal ribosome entry site (HCV IRES) domain IV in translation initiation and regulation, two chimeric IRES elements were constructed to contain the reciprocal domain IV in the otherwise HCV and classical swine fever virus IRES elements. This permitted an examination of the role of domain IV in the control of HCV translation. A specific inhibitor of the HCV IRES, vitamin B12, was shown to inhibit translation directed by all IRES elements which contained domain IV from the HCV and the GB virus B IRES elements, whereas the HCV core protein could only suppress translation from the wild-type HCV IRES. Thus, the mechanisms of translation inhibition by vitamin B12 and the core protein differ, and they target different regions of the IRES.

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