title: Autocatalytic activity of the ubiquitin-specific protease domain of herpes simplex virus 1 VP1-2.
abstract: The herpes simplex virus (HSV) tegument protein VP1-2 is essential for virus entry and assembly. VP1-2 also contains a highly conserved ubiquitin-specific protease (USP) domain within its N-terminal region. Despite conservation of the USP and the demonstration that it can act on artificial substrates such as polyubiquitin chains, identification of the relevance of the USP in vivo to levels or function of any substrate remains limited. Here we show that HSV VP1-2 USP can act on itself and is important for stability. VP1-2 N-terminal variants encompassing the core USP domain itself were not affected by mutation of the catalytic cysteine residue (C65). However, extending the N-terminal region resulted in protein species requiring USP activity for accumulation. In this context, C65A mutation resulted in a drastic reduction in protein levels which could be stabilized by proteosomal inhibition or by the presence of normal C65. The functional USP domain could increase abundance of unstable variants, indicating action at least in part, in trans. Interestingly, full-length variants containing the inactive USP, although unstable when expressed in isolation, were stabilized by virus infection. The catalytically inactive VP1-2 retained complementation activity of a VP1-2-negative virus. Furthermore, a recombinant virus expressing a C65A mutant VP1-2 exhibited little difference in single-step growth curves and the kinetics and abundance of VP1-2 or a number of test proteins. Despite the absence of a phenotype for these replication parameters, the USP activity of VP1-2 may be required for function, including its own stability, under certain circumstances.
authors.affiliation: Section of Virology, Faculty of Medicine, Imperial College London, St. Mary's Campus, Norfolk Place, London W2 1PG, United Kingdom.
authors.forename: M
authors.initials: M
authors.lastname: Bolstad
authors.forename: F
authors.initials: F
authors.lastname: Abaitua
authors.forename: C M
authors.initials: CM
authors.lastname: Crump
authors.forename: P
authors.initials: P
authors.lastname: O'Hare
mesh_terms: D000818:Animals; D002460:Cell Line; D004252:DNA Mutational Analysis; D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism*; D005816:Genetic Complementation Test; D018259:Herpesvirus 1, Human / Q000201:enzymology* / Q000254:growth & development; D006801:Humans; D064570:Ubiquitin-Specific Proteases; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: C078035:UL36 protein, Human herpesvirus 1; D014764:Viral Proteins; D010450:Endopeptidases; D064570:Ubiquitin-Specific Proteases