INITIATION OF HERPES-SIMPLEX VIRUS THYMIDINE KINASE POLYPEPTIDES

When employed as a transgene reporter, the herpes simplex type 1 virus (HSV1) thymidine kinase gene (tk) is ectopically expressed in mouse t estis, The principal testicular mRNA lacks the 5'-end of the tk readin g frame, As a result the principal translation products, P2 and P3, ar e N-terminally truncated. These co-migrate in SDS-PAGE with polypeptid es synthesised during HSV1 infection that were previously thought to b e initiated at methionine codons ATG(46) and ATG(60) Prompted by these observations we generated modified tk genes each carrying only one of the first three ATG codons. Transfected cells expressed both full-len gth enzyme (P1) and P2 when only ATG(1) was unmodified, P2 and P3 when only ATG(46) was unmodified or P2 and a fourth polypeptide (P4) when only ATG(60) was unmodified. Our observations indicate that P3 is init iated at ATG(46) rather than ATG(60), while P2 is initiated at a non-A TG codon rather than ATG(46) and Pq is initiated at ATG(60). When eith er of two putative non-ATG initiation codons was modified P2 was no lo nger produced, Cells mainly expressing either P1 or P3 exhibited the s ame sensitivity to Ganciclovir as cells transfected with the unaltered tk gene, P1 and P3 both have TK activity while P4 probably has none.