A BACULOVIRUS MUTANT DEFECTIVE IN PKIP, A PROTEIN WHICH INTERACTS WITH A VIRUS-ENCODED PROTEIN-KINASE

We have found that a temperature-sensitive mutant of the baculovirus A cMNPV, tsB97, is defective in PKIP, the product of ORF24 which was pre viously found to interact with and stimulate the activity of a virus-e ncoded protein kinase, PK-1. The mutant lacks the ability to form plaq ues and occlusion bodies at the ronpermissive temperature. The mutant displays several properties which suggest a defect in the latter half of the late phase of infection; these properties include a delay in th e shutoff of host protein synthesis, the presence of aberrant electron -dense bodies associated with the virogenic stroma, and the production of few, if any, progeny budded virus. A study of the expression of se lected late genes showed no difference in the timing or level of trans cription or translation of most late ger;es. However, elevated levels of the late 6.9K protein, a protamine-like protein, were observed in m utant-infected cells at 24 h postinfection, suggesting a defect in the regulation of this protein. Two polypeptides, 40 and 6 kDa, exhibited considerably higher levels of steady-state phosphorylation in wt-infe cted cells versus tsB97-infected cells at 24 h p.i. and could be candi dates for PK-1/PKIP-mediated phosphorylation. The tsB97 mutant also di splayed a severe defect in very late gene transcription which accounts for its inability to form occlusion bodies. The effect of PKIP on ver y late gene transcription may be a secondary effect of the block in th e late phase of infection. PKIP showed no ability to transactivate exp ression from a very late promoter in transient expression assays. (C) 1998 Academic Press.