The p10 gene of Bombyx mori nucleopolyhedrosis virus encodes a 7.5-kDa protein and is hypertranscribed from a TAAG motif

Abstract

In baculovirus-based high-level expression of cloned foreign genes, the viral very late gene promoters ofpolyhedrin (polh) andp10 are extensively exploited. Here we report the cloning and characterization of thep10 gene from a local isolate ofBombyx mori nucleopolyhedrosis virus (BmNPV). The gene harbours a 213-bp open reading frame encoding a protein of 70 amino acids with a predicted molecular mass of 7.5 kDa. The BmNPVp10 showed deletion of a single A at +210 nucleotide compared to the prototype baculovirus,Autographa californica multinucleocapsid nucleopolyhedrosis virus (AcMNPV),p10 gene, resulting in a translational frameshift to generate a termination codon and consequently a truncated polypeptide instead of the 10-kDa protein. This protein P7.5 from BmNPV has a putative leucine zipper dimerization motif towards the N-terminal end and the central nuclear disintegration domain but the carboxy-terminal domain implicated in protein association for fibrillar structure formation is absent. Phylogenetic analysis revealed thatp10 is highly conserved among baculoviruses and the BmNPV strains are more closely related to AcMNPV than other baculoviruses. The transcription ofp10 is regulated in a temporal manner, reaching maximal levels by 72 h post-infection. RNAase protection and primer extension analysis mapped the transcription start sites at -70 and-71 nt with respect to the ATG, within the conserved baculovirus late gene motif TAAG. The upstream region showed complete homology to the strong promoter of the AcMNPVp10, suggesting that this promoter from BmNPV could also be exploited for high-level expression of cloned foreign genes in silkworm cells or larvae.