Identification of a highly conserved, functional nuclear localization signal within the N-terminal region of herpes simplex virus type 1 VP1-2 tegument protein.
VP1-2 is a large structural protein assembled into the tegument compartment of the virion, conserved across the herpesviridae, and essential for virus replication. In herpes simplex virus (HSV) and pseudorabies virus, VP1-2 is tightly associated with the capsid. Studies of its assembly and function remain incomplete, although recent data indicate that in HSV, VP1-2 is recruited onto capsids in the nucleus, with this being required for subsequent recruitment of additional structural proteins. Here we have developed an antibody to characterize VP1-2 localization, observing the protein in both cytoplasmic and nuclear compartments, frequently in clusters in both locations. Within the nucleus, a subpopulation of VP1-2 colocalized with VP26 and VP5, though VP1-2-positive foci devoid of these components were observed. We note a highly conserved basic motif adjacent to the previously identified N-terminal ubiquitin hydrolase domain (DUB). The DUB domain in isolation exhibited no specific localization, but when extended to include the adjacent motif, it efficiently accumulated in the nucleus. Transfer of the isolated motif to a test protein, beta-galactosidase, conferred specific nuclear localization. Substitution of a single amino acid within the motif abolished the nuclear localization function. Deletion of the motif from intact VP1-2 abrogated its nuclear localization. Moreover, in a functional assay examining the ability of VP1-2 to complement growth of a VP1-2-ve mutant, deletion of the nuclear localization signal abolished complementation. The nuclear localization signal may be involved in transport of VP1-2 early in infection or to late assembly sites within the nucleus or, considering the potential existence of VP1-2 cleavage products, in selective localization of subdomains to different compartments.
Abaitua
F
F
Marie Curie Research Institute, The Chart, Oxted, Surrey RH8 0TL, United Kingdom.
O'Hare
P
P
D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D017124:Conserved Sequence; D017353:Gene Deletion; D018259:Herpesvirus 1, Human / Q000737:chemistry* / Q000378:metabolism*; D008969:Molecular Sequence Data; D019913:Nuclear Localization Signals; D016415:Sequence Alignment; D014764:Viral Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
D019913:Nuclear Localization Signals; D014764:Viral Proteins