US3 kinase-mediated phosphorylation of tegument protein VP8 plays a critical role in the cellular localization of VP8 and its effect on the lipid metabolism of bovine herpesvirus-1-infected cells.

Bovine herpesvirus-1 (BoHV-1) infects bovine species, causing respiratory infections, genital disorders and abortions. VP8 is the most abundant tegument protein of BoHV-1 and critical for virus replication in cattle. In this study the cellular transport of VP8 in BoHV-1-infected cells and its ability to alter the cellular lipid metabolism was investigated. A viral kinase, US3, was found to be involved in regulating these processes. In the early stages of infection VP8 was localized in the nucleus. Subsequently, presumably after completion of its role in the nucleus, VP8 was translocated to the cytoplasm. When US3 was deleted or the essential US3-phosphorylation site of VP8 was mutated in BoHV-1, the majority of VP8 was localized in the nucleus of infected cells. This suggests that phosphorylation by US3 may be critical for cytoplasmic localization of VP8. Eventually, the cytoplasmic VP8 was accumulated in the cis-Golgi apparatus, but not in the trans-Golgi network, implying that VP8 was not involved in virion transport towards and budding from the cell membrane. VP8 caused lipid droplet (LD) formation in the nucleus of transfected cells and increased cellular cholesterol levels. Lipid droplets were not found in the nucleus of BoHV-1-infected cells, when VP8 is cytoplasmic in the presence of US3. However, when US3 was deleted or phosphorylation residues in VP8 were mutated, nuclear VP8 and LDs appeared in BoHV-1-infected cells. The total cholesterol level was increased in BoHV-1-infected cells, but not in ΔUL47-BoHV-1-infected cells further supporting a role for VP8 in altering the cellular lipid metabolism during infection. [ABSTRACT FROM AUTHOR]