IDENTIFICATION OF A VIRAL GENE ENCODING A UBIQUITIN-LIKE PROTEIN

The baculovirus Autographa californica nuclear polyhedrosis virus (AcMNPV, which is representative of the MNPV subtype in which the virions may contain many nucleocapsids within a single viral envelope) encodes a protein, v-ubi, that has 76% identity with the eukaryotic protein ubiquitin. Transcription mapping indicated that the gene for v-ubi was transcribed during the late phase of viral infection. Two transcription start sites potentially encoding v-ubi were identified. Both sites were contained within a sequence motif common to baculovirus late genes. A recombinant virus, AcUbi-βGal, encoding a ubiquitin-β-galactosidase fusion protein was constructed to monitor the temporal regulation of v-ubi gene during viral infection. The fusion protein was expressed maximally at 14-18 hr postinfection, consistent with its classification as a late protein. The amount of ubiquitin-β-galactosidase fusion protein that accumulated in AcUbi-βGal-infected cells by 48 hr postinfection was ~ 14% of the level of β-galactosidase that was synthesized under control of the polyhedrin promoter. Transcriptional analysis confirmed that synthesis of the fusion protein was directed by the v-ubi gene promoter. AcUbi-βGal also produced normal levels of authentic viral ubiquitin message. Southern blot analysis of AcUbi-βGal and 15 additional isolates revealed that the fusion sequences had not recombined at the ubiquitin locus. A polyubiquitin gene was isolated and sequenced from Spodoptera frugiperda, a lepidopteran host cell line for AcMNPV. The predicted amino acid sequence of the product of the host gene is identical to animal ubiquitin.