Expression kinetics of the late UL12 gene encoding the bovine herpesvirus 1 alkaline nuclease

We characterized the expression kinetics of the transcript and protein gene rated from the bovine herpesvirus 1 (BHV1) homologue of the herpes simplex virus 1 (HSV1) UL12 gene that encodes a viral alkaline nuclease. The BHV1 U L12 coding sequence, which was previously shown to express in E. coli a pro tein exhibiting nuclease activity, is located at positions 82776 --> 84239 of the viral genome. Northern blot analysis of RNA from BHV1-infected cells with a single strand RNA probe complementary to UL12 detected four specifi c 3' coterminal viral transcripts of 4.2, 3.7, 2.2, and 0.7 kb that accumul ated simultaneously from 6 to 24 hours post-infection (p.i.). S1 nuclease m apping of the UL12 capping site at position 82384 of the genome as well as the identification of a consensus polyadenylation signal at 84490-84495 all owed us to establish that the 2.2kb transcript corresponds to that of UL12. A UL12 specific antiserum generated against a T7-Tag/UL12 fusion protein e xpressed in E. coli detected a 53 kDa protein in cell lysates from BHV1-inf ected cells, whose size correlated with that predicted (51,844 Da), which a ccumulated from 12 to 30 h p.i. Differences observed between the transcript ional and translational expression profiles suggest that the UL12 of BHV1 i s regulated at both the translational and posttranslational levels. Surpris ingly, the protein expression was strictly dependent on viral DNA synthesis , unambiguously demonstrating that BHV1 UL12 belongs to viral arenes of the gamma2 class. This is in contrast to the HSV1 and pseudorabies homologues that are classified as early (beta) genes. Further studies will be required to determine whether these kinetic differences have any functional implica tions.