Characterization of the ecdysteroid UDP-Glucosyltransferase (egt) gene of Anticarsia gemmatalis nucleopolyhedrovirus

The Anticarsia gemmatalis nucelopolyhedrovirus (AgMNPV) egt gene was cloned , sequenced and its expression characterized by RT-PCR and western blot ana lysis. Sequence analysis of the gene indicated the presence of an open read ing frame (ORF) of 1482 nucleotides, which codes for a polypeptide of 494 a mino acids. A TATA box and a conserved regulatory sequence (CATT) found in other baculovirus early genes were present in the promoter region of the eg t gene. A poly-A consensus sequence was present in the 3' untranslated regi on (3'-UTR) of the gene. Homology comparisons showed that the EGT protein o f AgMNPV is most closely related (95.9% amino acid sequence identity) to th e EGT from the Choristoneura fumiferana DEF nucleopolyhedrovirus (CfDEF). T ranscriptional analysis of the AgMNPV egt gene showed that egt-specific tra nscripts can be detected both early and late in infection. The EGT protein was detected, by western blot analysis, in the intra- (from 12 to 48 h post -infection) and extra-cellular (from 12 to 96 h post-infection) fractions o f infected insect cells. The AgMNPV Bgl II-F fragment, which has homology t o the AcMNPV ie-1 gene, was cloned and used to cotransfect SF21 cells with the cloned AgMNPV egt gene. EGT activity was observed, suggesting that AgMN PV ie-1 can transactivate egt expression.