Cleavage of the hepatitis C virus NS5A protein by caspase-3 in the interferon sensitivity-determining region in a sequence-dependent manner.

Hepatitis C virus (HCV) nonstructural protein 5A (NS5A) has versatile functions and has been implicated in viral pathogenesis, including interferon (IFN) resistance and hepatocarcinogenesis. It has been reported that NS5A is cleaved into a few fragments by a caspase(s) or caspase-like enzyme(s) under certain conditions. Two cleavage sites have been mapped to the Asp residues at positions 154 and 398 (D154 and D398). However, other possible cleavage sites were not determined yet so far. In this study, we demonstrated caspase-3-mediated NS5A cleavage upon apoptotic stimuli and identified a new site as the third cleavage target. This site was mapped to D251, which lies within IFN sensitivity-determining region (ISDR). Although D251 was conserved among all HCV subtype 1b (HCV-1b) strains tested, the consensus caspase-3 recognition sequence (D248-X-X-D251) was not conserved due to the sequence variation at position 248 in ISDR. Furthermore, A252 was found to be necessary for efficient cleavage of NS5A. The virological significance of the HCV strain-dependent NS5A cleavage at this site awaits further investigation.