The Hepatitis E Virus Open Reading Frame 3 Protein Activates ERK through Binding and Inhibition of the MAPK Phosphatase

Kar-Roy, Anindita; Korkaya, Hasan; Oberoi, Ruchi; Lal, Sunil K.; Jameel, Shahid

doi:10.1074/jbc.m400457200

Cited by 79 publications

(89 citation statements)

References 69 publications

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“…This suggests that there may be further, as yet unidentified, ways in which NS5A and/or other nonstructural proteins are able to modulate the Ras-ERK pathway. In this regard, the hepatitis E virus ORF3 protein was recently shown to activate ERK by inactivating an ERK-specific phosphatase (Kar-Roy et al, 2004). It is intriguing to speculate that, as in the case of the EGFR and receptor-proximal events in the EGFR signalling pathway.…”

Section: Discussionmentioning

confidence: 99%

Perturbation of epidermal growth factor receptor complex formation and Ras signalling in cells harbouring the hepatitis C virus subgenomic replicon

Macdonald

Chan

Harris

2005

Journal of General Virology

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Hepatitis C virus non-structural NS5A protein inhibits epidermal growth factor (EGF)-stimulated activation of the Ras-ERK mitogen-activated protein kinase pathway at a point upstream of Ras activation. To determine the mechanism of this inhibition, the events occurring between the EGF receptor and Ras in Huh-7 cells harbouring the HCV subgenomic replicon were investigated. It was shown that, following EGF stimulation, these cells exhibited decreased EGF receptor tyrosine phosphorylation, aberrant recruitment of the adaptor proteins ShcA and Grb2 to the EGF receptor, reduced phosphorylation of ShcA and reduced Ras activation in comparison with control cells. These data are consistent with effects of NS5A and/or other components of the replicon on multiple events occurring upstream of Ras. INTRODUCTIONHepatitis C virus (HCV) is the leading cause of liver disease and transplantation in the developing world and is estimated to infect 3 % of the global population (WHO, 1999). More than 80 % of those infected will develop a chronic disease culminating in liver fibrosis, cirrhosis and hepatocellular carcinoma. The HCV genome is a 9?5 kb positive-sense RNA molecule that is translated in a capindependent fashion via an internal ribosome entry site to generate a 3000 residue polypeptide, cleaved by host and viral proteases to produce 10 mature viral proteins.The NS5A protein is one of six non-structural proteins that are believed to form a membrane-bound RNA-replication complex . Further to this role, NS5A has been shown to modulate a range of cellular signalling pathways, including the mitogen-activated protein kinase (MAPK) pathways. In particular, we and others have shown that NS5A expression is able to inhibit the Ras-ERK MAPK pathway (Tan et al., 1999;Georgopoulou et al., 2003;Macdonald et al., 2003). This pathway is activated by growth factors binding to their cognate receptors; this induces autophosphorylation of the receptors on tyrosine residues, which then act as ligands for the Src homology 2 (SH2) domains of adaptor proteins, such as ShcA and Grb2. Grb2 associates with a guanine nucleotideexchange factor, Sos, which induces GDP/GTP exchange on the GTP-binding protein Ras (Tari & Lopez-Berestein, 2001). Ras then binds to and activates a serine kinase, Raf, which triggers a kinase cascade ultimately leading to c-Fos expression and concomitant activation of the AP-1 transcription factor (of which c-Fos is a component). We previously demonstrated that transfection of a dominant-active mutant of Ras was able to override the NS5A-mediated block to this pathway and we therefore concluded that NS5A was acting between the epidermal growth factor receptor (EGFR) and the activation of Ras (Macdonald et al., 2003). The mechanism of this inhibition is unknown, although it has been shown that NS5A is able to bind Grb2 (Tan et al., 1999;, By doing so, it is possible that NS5A might inhibit the formation of the Grb2-Sos complex, thereby breaking the link between the activated EGFR and Ras. However, it has previously been sta...

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Section: Discussionmentioning

confidence: 99%

Perturbation of epidermal growth factor receptor complex formation and Ras signalling in cells harbouring the hepatitis C virus subgenomic replicon

Macdonald

Chan

Harris

2005

Journal of General Virology

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“…ORF3, mapped between ORF1 and ORF2, encodes a 13.5-kDa protein that is associated with the membrane as well as with the cytoskeleton fraction (27). This protein is shown to be phosphorylated by the cellular mitogen-activated protein kinase (6,8). The ORF3 protein may have a regulatory function (6,8).…”

Section: Hepatitis E Virus (Hev) Is a Noncultivable Virus That Causesmentioning

confidence: 99%

“…This protein is shown to be phosphorylated by the cellular mitogen-activated protein kinase (6,8). The ORF3 protein may have a regulatory function (6,8). Ever since HEV was first discovered in 1980 and visualized by immune electron microscopy in 1983 (2), many efforts have been made, using different expression systems, to express the structural protein (5,11,17,26).…”

Section: Hepatitis E Virus (Hev) Is a Noncultivable Virus That Causesmentioning

confidence: 99%

Essential Elements of the Capsid Protein for Self-Assembly into Empty Virus-Like Particles of Hepatitis E Virus

Takeda

Miyamura

et al. 2005

J Virol

166

205

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“…The exact functions of vp13 in HEV infection remain unknown although the findings of a number of studies have shown that it plays a role in cellular signaling pathways (13,17,24,(34)(35)(36)40). During subcellular fractionation of COS-7 cells transfected with a vp13-expressing plasmid, vp13 was found to partition with the cytoskeletal fraction (40).…”

Section: Hepatitis E Virus (Hev) Is the Causative Agent Of Hepatitis mentioning

confidence: 99%

The Hepatitis E Virus Open Reading Frame 3 Product Interacts with Microtubules and Interferes with Their Dynamics

Kannan

Fan

Patel

et al. 2009

J Virol

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Hepatitis E virus (HEV) is the causative agent of hepatitis E, a major form of viral hepatitis in developingcountries. The open reading frame 3 (ORF3) of HEV encodes a phosphoprotein with a molecular mass of approximately 13 kDa (hereinafter called vp13). vp13 is essential for establishing HEV infections in animals, yet its exact functions are still obscure. Our current study found evidence showing interaction between vp13 and microtubules. Live-cell confocal fluorescence microscopy revealed both filamentous and punctate distribution patterns of vp13 in cells transfected with recombinant ORF3 reporter plasmids. The filamentous pattern of vp13 was altered by a microtubule-destabilizing drug. The vp13 expression led to elevation of acetylated ␣-tubulin, indicating increased microtubule stability. Its association with microtubules was further supported by its presence in microtubule-containing pellets in microtubule isolation assays. Exposure of these pellets to a high-salt buffer caused release of the vp13 to the supernatant, suggesting an electrostatic interaction. Inclusion of ATP and GTP in the lysis buffer during microtubule isolation also disrupted the interaction, indicating its sensitivity to the nucleotides. Further assays showed that motor proteins are needed for the vp13 association with the microtubules because disruption of dynein function abolished the vp13 filamentous pattern. Analysis of ORF3 deletion constructs found that both of the N-terminal hydrophobic domains of vp13 are needed for the interaction. Thus, our findings suggest that the vp13 interaction with microtubules might be needed for establishment of an HEV infection.The hepatitis E virus (HEV), the sole member of the genus Hepevirus, is a single-strand positive-sense RNA virus that is the causative agent in endemics and epidemics of acute human hepatitis in many parts of the world (5). Transmitted mainly from contaminated water through the fecal-oral route, HEV infection causes a fulminant form of hepatitis that has a mortality rate of up to 20% in pregnant women (28). HEV infection is considered zoonotic. Swine and chicken HEV strains have been found in the United States (11,23). A swine strain can infect chimpanzees under experimental conditions, and a human strain that is genetically similar to the swine strain can experimentally infect pigs (22). Direct evidence of the zoonotic nature of HEV infection has been provided in reports of a series of cases of HEV infection in people who ate undercooked deer meat 6 to 7 weeks before the onset of the disease (19,33,39). HEV RNA recovered from the leftover deer meat was found to be identical in nucleotide sequence to the HEV RNA recovered from the individuals who became ill (31).The HEV genome is approximately 7.2 kb in length and consists of three open reading frames (ORFs) (32). ORF1 encodes a nonstructural polyprotein that includes the RNAdependent RNA polymerase. ORF2 encodes the capsid protein, the major structural protein in virion. ORF3 encodes a phosphoprotein that was found to be esse...

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The Hepatitis E Virus Open Reading Frame 3 Protein Activates ERK through Binding and Inhibition of the MAPK Phosphatase

Cited by 79 publications

References 69 publications

Perturbation of epidermal growth factor receptor complex formation and Ras signalling in cells harbouring the hepatitis C virus subgenomic replicon

Perturbation of epidermal growth factor receptor complex formation and Ras signalling in cells harbouring the hepatitis C virus subgenomic replicon

Essential Elements of the Capsid Protein for Self-Assembly into Empty Virus-Like Particles of Hepatitis E Virus

The Hepatitis E Virus Open Reading Frame 3 Product Interacts with Microtubules and Interferes with Their Dynamics

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