THE P95 GENE OF BOMBYX-MORI NUCLEAR POLYHEDROSIS-VIRUS - TEMPORAL EXPRESSION AND FUNCTIONAL-PROPERTIES

As part of our effort to identify baculovirus proteins acting as trans criptional regulators, we have characterized a gene, p95, of Bombyx mo ri nuclear polyhedrosis virus (BmNPV) that encompasses an open reading frame for a putative 95-kDa polypeptide (P95), The N-terminal half of the conceptually translated P95 contains two zinc finger-type DNA-bin ding motifs, and its C terminus contains a proline-rich region reminis cent of transcriptional activation regions. Northern blot analysis ind icates that two mRNA species, 3.5 and 1.7 kb in size, are transcribed from the p95 gene at different times postinfection, These two mRNA spe cies are produced by differential polyadenylation site usage. While th e longer transcript can encode the P95 protein, the shorter one may en code a prematurely terminated version of the P95 polypeptide produced by ribosome frameshifting occurring at heptanucleotide ''slippage') si tes located near the relevant polyadenylation site. Transcription of t he p95 gene is initiated at a proximal site located 70 nucleotides ups tream of the translation start codon of P95, a middle site located 170 nucleotides from the start codon, and a set of three closely spaced d istal sites located 385, 390, and 409 nucleotides from the translation start codon, The middle and distant initiation sites are utilized bef ore and after BmNPV DNA replication, while transcripts initiated at th e proximal site occur largely during the late and very late stages of viral infection. Transient-expression assays indicate that P95 can sti mulate gene expression driven by the promoter of its own gene and the promoter of the cytoplasmic actin gene of B. mori, The P95-mediated tr ans activation can be further augmented by BmIE1, an immediate early g ene product of BmNPV, In contrast to the case with the actin promoter, however, the promoter of the p95 gene can be trans activated by the p roduct of its own gene only in the presence of BmIE1, Our data suggest that proteins P95 and BmIE1 of BmNPV and, by analogy, those of other baculoviruses may interact with each other and synergize to potentiate transcription.