IDENTIFICATION AND CHARACTERIZATION OF THE SPODOPTERA-LITTORALIS NUCLEOPOLYHEDROVIRUS TYPE-B LEF-3 GENE

We have identified a gene from the Spodoptera littoralis nucleopoly-he drovirus type B (SpliNPV-B) with several characteristics that suggest that it is homologous to the lef-3 genes of the Autographa californica and Orgyia pseudotsugata NPVs (AcMNPV and OpMNPV, respectively). The SpliNPV-B lef-3 gene was mapped between 43.6 and 45.5 map units of the SpliNPV-B genome. Northern blot analysis showed that SpliNPV-B lef-3 was expressed as a 1.6 Kb transcript at 5 h post infection (p.i.), rea ched high levels at 24 h p.i., and remained highly expressed at 56 h p .i. Transcription of SpliNPV-B lef-3 initiated at two distinct sites d ownstream from a TATA-box motif and terminated 25 nucleotides downstre am from the translation stop site of the putative LEF-3 polypeptide. T he 5'-boundaries of lef-3 promoter elements were investigated by trans ient expression assays, which revealed that the major components of th e lef-3 promoter are within a 183 base pair region upstream of the dis tal transcription initiation site. Transfection of SpliNPV-B infected Sf9 cells with anti-sense oligonucleotides designed to inhibit LEF-3 e xpression resulted in substantial reduction of viral DNA replication, suggesting that the role of SpliNPV-B lef-3 may be similar to that of AcMNPV and OpMNPV lef-3 genes, which are essential for viral DNA repli cation.