Hepatitis C Virus Nonstructural Proteins Are Localized in a Modified Endoplasmic Reticulum of Cells Expressing Viral Subgenomic Replicons

Mottola, Giovanna; Cardinali, Giorgia; Ceccacci, Alessandra; Trozzi, Caterina; Bartholomew, Linda; Torrisi, Maria Rosaria; Pedrazzini, Emanuela; Bonatti, Stefano; Migliaccio, Giovanni

doi:10.1006/viro.2001.1229

Cited by 155 publications

(157 citation statements)

References 44 publications

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“…The immunohistochemical staining showed granular cytoplasmic signals, consistent with synthesis in association with the endoplasmic reticulin networks. 18 These experiments confirm that FCA1 replicon system exhibits efficient viral RNA replication and generates abundant viral nonstructural proteins.…”

Section: Resultssupporting

confidence: 62%

Gene Expression Associated With Interferon Alfa Antiviral Activity in An Hcv Replicon Cell Line

Zhu¹,

Zhao²,

Collins

et al. 2003

Hepatology

100

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Section: Resultssupporting

confidence: 62%

Gene Expression Associated With Interferon Alfa Antiviral Activity in An Hcv Replicon Cell Line

Zhu¹,

Zhao²,

Collins

et al. 2003

Hepatology

100

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“…When expressed either alone, or in conjunction with other HCV proteins in the context of the subgenomic replicon, HCV NS5A displays a punctate cytoplasmic staining that is at least partially associated with the endoplasmic reticulum (Mottola et al, 2002; McCormick et al, 2006). Both HCV and GBV-B NS5A proteins contain an N-terminal amphipathic helix reported to mediate membrane association (Brass et al, 2002(Brass et al, , 2007.…”

Section: Resultsmentioning

confidence: 99%

“…Much work has focused on the HCV NS5A protein, it is believed to be a component of an RNA replication complex on cytoplasmic membranes together with the other nonstructural proteins (Egger et al, 2002;Lohmann et al, 1999;Mottola et al, 2002). In addition NS5A has been shown to bind to and modulate a range of cellular proteins, influencing activation of signalling pathways .…”

Section: Introductionmentioning

confidence: 99%

“…There are three structural proteins at the N terminus: core, E1 and E2, followed by a small cation channel, p7 (p13 in GBV-B). The C-terminal two-thirds of the polyprotein comprises six non-structural proteins: NS2, NS3, NS4A, NS4B, NS5A and NS5B.Much work has focused on the HCV NS5A protein, it is believed to be a component of an RNA replication complex on cytoplasmic membranes together with the other nonstructural proteins (Egger et al, 2002;Lohmann et al, 1999;Mottola et al, 2002). In addition NS5A has been shown to bind to and modulate a range of cellular proteins, influencing activation of signalling pathways .…”

mentioning

confidence: 99%

“…In common with HCV NS5A, both proteins exhibited a significantly larger apparent molecular mass in comparison with their actual molecular mass, for FLAG-tagged GBV-B NS5A this value is 45 kDa. This discrepancy may be due to the high proline content of the protein (10 %) or to post-translational modifications.The subcellular distribution of the GBV-B NS5A protein differs from that of the HCV NS5A proteinWhen expressed either alone, or in conjunction with other HCV proteins in the context of the subgenomic replicon, HCV NS5A displays a punctate cytoplasmic staining that is at least partially associated with the endoplasmic reticulum (Mottola et al, 2002; McCormick et al, 2006). Both HCV and GBV-B NS5A proteins contain an N-terminal amphipathic helix reported to mediate membrane association (Brass et al, 2002(Brass et al, , 2007.…”

mentioning

confidence: 99%

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A comparative cell biological analysis reveals only limited functional homology between the NS5A proteins of hepatitis C virus and GB virus B

Mankouri

Milward

Pryde

et al. 2008

Journal of General Virology

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GB virus B (GBV-B) is the closest relative to hepatitis C virus (HCV) with which it shares a common genome organization, however, unlike HCV in humans, it generally causes an acute resolving hepatitis in New World monkeys. It is important to understand the factors regulating the different disease profiles of the two viruses and in this regard, as well as playing a key role in viral RNA replication, the HCV NS5A non-structural protein modulates a variety of host-cell signalling pathways. We have shown previously that HCV NS5A, expressed either alone, or in the context of the complete polyprotein, inhibits the Ras-extracellular-signal-regulated kinase (Erk) pathway and activates the phosphoinositide 3-kinase (PI3K) pathway. In this report, we investigate whether these functions are shared by GBV-B NS5A. Immunofluorescence analysis revealed that a C-terminally FLAG-tagged GBV-B NS5A exhibited a punctate cytoplasmic distribution. However, unlike HCV NS5A, the GBV-B protein did not partially co-localize with early endosomes. Utilizing a transient luciferase reporter system, we observed that GBV-B NS5A failed to inhibit Ras-Erk signalling, however GBV-B NS5A expression did result in the elevation of b-catenin-dependent transcription via activation of the PI3K pathway. These effects of GBV-B and HCV NS5A on the PI3K and Ras-Erk pathways were confirmed in cells harbouring subgenomic replicons derived from the two viruses. Based on these data we speculate that the differential effects of the two NS5A proteins on cellular signalling pathways may contribute to the differences in the natural history of the two viruses. INTRODUCTIONGB virus B (GBV-B) is the closest phylogenetic relative of hepatitis C virus (HCV) (Muerhoff et al., 1995;Ohba et al., 1996); it shares up to 28 % overall amino acid identity with HCV, rising to 50 % conservation in regions such as the NS3 coding sequence. GBV-B has been proposed as a model for HCV infection and has been used to characterize candidate antivirals as well as for pathogenesis studies (Beames et al., 2001;Bright et al., 2004). It shares the hepatotropism of HCV. Although GBV-B has been reported to lead to persistent infections in some cases in tamarins (Saguinus species) (Martin et al., 2003;Nam et al., 2004), unlike the human virus, GBV-B primarily causes an acute, resolving hepatitis in tamarins and other New World monkeys such as common marmosets (Callithrix jacchus) (Bukh et al., 2001;Jacob et al., 2004;Lanford et al., 2003). In contrast, HCV is a major cause of chronic hepatitis, liver cirrhosis and hepatocellular carcinoma, affecting an estimated 3 % of the world's population.Both GBV-B and HCV are members of the genus Hepacivirus of the family Flaviviridae and have a positivesense RNA genome that is translated in a cap-independent fashion to generate a polyprotein, cleaved by host and viral proteases to produce 10 mature viral proteins. There are three structural proteins at the N terminus: core, E1 and E2, followed by a small cation channel, p7 (p13 in GBV-B). The C-terminal t...

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Structure and Molecular Virology

McGarvey¹,

Houghton²

2005

Viral Hepatitis

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Hepatitis C Virus Nonstructural Proteins Are Localized in a Modified Endoplasmic Reticulum of Cells Expressing Viral Subgenomic Replicons

Cited by 155 publications

References 44 publications

Gene Expression Associated With Interferon Alfa Antiviral Activity in An Hcv Replicon Cell Line

Gene Expression Associated With Interferon Alfa Antiviral Activity in An Hcv Replicon Cell Line

A comparative cell biological analysis reveals only limited functional homology between the NS5A proteins of hepatitis C virus and GB virus B

Structure and Molecular Virology

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