The inhibitor of apoptosis gene (iap-3) of Anticarsia gemmatalis multicapsid nucleopolyhedrovirus (AgMNPV) encodes a functional IAP.

Programmed cell death or apoptosis is one of the defense mechanisms used by insect cells in response to baculovirus infection. Baculoviruses harbour antiapoptotic genes to prevent apoptosis and to maintain the normal course of infection. In this work, we showed that, like other baculoviruses, Anticarsia gemmatalis multicapsid nucleopolyhedrovirus (AgMNPV) has a functional inhibitor of apoptosis gene (iap-3). The iap-3 gene was cloned, sequenced and its transcription confirmed by RT-PCR. The putative iap-3 gene of the baculovirus AgMNPV has 864 nucleotides and codes an ORF of 287 amino acids. We have found two BIR motifs (baculoviral iap repeats) at the amino-terminal region and a carboxi-terminal RING finger motif. The IAP-3 protein of AgMNPV is closely related to IAP-3 proteins of baculoviruses and lepidopteran IAPs, with most amino acid identity (75%) with the IAP-3 protein of CfDefNPV (Choristoneura fumiferana DEF nucleopolyhedrovirus). Transcriptional analysis of the AgMNPV iap-3 gene showed that iap-3-specific transcripts could be detected early and late in the infection. The iap-3 gene of AgMNPV was shown to encode a functional IAP since insect cells transfected with increasing amounts of a plasmid containing the iap-3 of AgMNPV showed increased resistance to apoptosis induced by a AgMNPV mutant virus.

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 analysis. Sequence analysis of the gene indicated the presence of an open reading frame (ORF) of 1482 nucleotides, which codes for a polypeptide of 494 amino acids. ATATA box and a conserved regulatory sequence (CATT) found in other baculovirus early genes were present in the promoter region of the egt gene. A poly-A consensus sequence was present in the 3' untranslated region (3'-UTR) of the gene. Homology comparisons showed that the EGT protein of AgMNPV is most closely related (95.9% amino acid sequence identity) to the EGT from the Choristoneura fumiferana DEF nucleopolyhedrovirus (CfDEF). Transcriptional analysis of the AgMNPV egt gene showed that egt-specific transcripts 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 of infected insect cells. The AgMNPV Bgl II-F fragment, which has homology to 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 AgMNPV ie-1 can transactivate egt expression.