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The lipid kinase phosphatidylinositol-4 kinase III alpha regulates the phosphorylation status of hepatitis C virus NS5A.


ABSTRACT: The lipid kinase phosphatidylinositol 4-kinase III alpha (PI4KIII?) is an essential host factor of hepatitis C virus (HCV) replication. PI4KIII? catalyzes the synthesis of phosphatidylinositol 4-phosphate (PI4P) accumulating in HCV replicating cells due to enzyme activation resulting from its interaction with nonstructural protein 5A (NS5A). This study describes the interaction between PI4KIII? and NS5A and its mechanistic role in viral RNA replication. We mapped the NS5A sequence involved in PI4KIII? interaction to the carboxyterminal end of domain 1 and identified a highly conserved PI4KIII? functional interaction site (PFIS) encompassing seven amino acids, which are essential for viral RNA replication. Mutations within this region were also impaired in NS5A-PI4KIII? binding, reduced PI4P levels and altered the morphology of viral replication sites, reminiscent to the phenotype observed by silencing of PI4KIII?. Interestingly, abrogation of RNA replication caused by mutations in the PFIS correlated with increased levels of hyperphosphorylated NS5A (p58), indicating that PI4KIII? affects the phosphorylation status of NS5A. RNAi-mediated knockdown of PI4KIII? or pharmacological ablation of kinase activity led to a relative increase of p58. In contrast, overexpression of enzymatically active PI4KIII? increased relative abundance of basally phosphorylated NS5A (p56). PI4KIII? therefore regulates the phosphorylation status of NS5A and viral RNA replication by favoring p56 or repressing p58 synthesis. Replication deficiencies of PFIS mutants in NS5A could not be rescued by increasing PI4P levels, but by supplying functional NS5A, supporting an essential role of PI4KIII? in HCV replication regulating NS5A phosphorylation, thereby modulating the morphology of viral replication sites. In conclusion, we demonstrate that PI4KIII? activity affects the NS5A phosphorylation status. Our results highlight the importance of PI4KIII? in the morphogenesis of viral replication sites and its regulation by facilitating p56 synthesis.

SUBMITTER: Reiss S 

PROVIDER: S-EPMC3649985 | biostudies-literature | 2013 May

REPOSITORIES: biostudies-literature

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The lipid kinase phosphatidylinositol-4 kinase III alpha regulates the phosphorylation status of hepatitis C virus NS5A.

Reiss Simon S   Harak Christian C   Romero-Brey Inés I   Radujkovic Danijela D   Klein Rahel R   Ruggieri Alessia A   Rebhan Ilka I   Bartenschlager Ralf R   Lohmann Volker V  

PLoS pathogens 20130509 5


The lipid kinase phosphatidylinositol 4-kinase III alpha (PI4KIIIα) is an essential host factor of hepatitis C virus (HCV) replication. PI4KIIIα catalyzes the synthesis of phosphatidylinositol 4-phosphate (PI4P) accumulating in HCV replicating cells due to enzyme activation resulting from its interaction with nonstructural protein 5A (NS5A). This study describes the interaction between PI4KIIIα and NS5A and its mechanistic role in viral RNA replication. We mapped the NS5A sequence involved in PI  ...[more]

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