Abstract
Like other viruses, productive hepatitis C virus (HCV) infection depends on certain critical host factors. We have recently shown that an interaction between HCV nonstructural protein NS5A and a host protein, TBC1D20, is necessary for efficient HCV replication. TBC1D20 contains a TBC (Tre-2, Bub2, and Cdc16) domain present in most known Rab GTPase-activating proteins (GAPs). The latter are master regulators of vesicular membrane transport, as they control the activity of membrane-associated Rab proteins. To better understand the role of the NS5A-TBC1D20 interaction in the HCV life cycle, we used a biochemical screen to identify the TBC1D20 Rab substrate. TBC1D20 was found to be the first known GAP for Rab1, which is implicated in the regulation of anterograde traffic between the endoplasmic reticulum and the Golgi complex. Mutation of amino acids implicated in Rab GTPase activation by other TBC domain-containing GAPs abrogated the ability of TBC1D20 to activate Rab1 GTPase. Overexpression of TBC1D20 blocked the transport of exogenous vesicular stomatitis virus G protein from the endoplasmic reticulum, validating the involvement of TBC1D20 in this pathway. Rab1 depletion significantly decreased HCV RNA levels, suggesting a role for Rab1 in HCV replication. These results highlight a novel mechanism by which viruses can hijack host cell machinery and suggest an attractive model whereby the NS5A-TBC1D20 interaction may promote viral membrane-associated RNA replication.
Highlights
Hepatitis C is a major global public health problem
The NS5A amphipathic ␣-helix is composed of a hydrophobic face proposed to be embedded in the cytoplasmic leaflet of the endoplasmic reticulum (ER) membrane and a polar charged face that is exposed to the cytosol and thought to mediate specific protein-protein interactions that are essential for the formation of a functional hepatitis C virus (HCV) replication complex [11]
We present here experiments that identify TBC1D20 as the Rab GTPase-activating proteins (GAPs) for Rab1 and suggest an attractive mechanism used by HCV to subvert host cell machinery for the promotion of viral replication
Summary
We have recently discovered the first example of a Rab GAP being exploited by a virus [21]. Starting from a yeast two-hybrid screen, HCV NS5A was shown to interact with the host cell protein TBC1D20 [21]. The latter contains a TBC domain present in most known Rab GAPs [22]. Depletion of TBC1D20 severely impairs HCV replication and prevents new infection, with minimal effects on cell viability [21]. These exciting results raised several key questions regarding the physiological role of TBC1D20 in host cells and in viral replication. We present here experiments that identify TBC1D20 as the Rab GAP for Rab and suggest an attractive mechanism used by HCV to subvert host cell machinery for the promotion of viral replication
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