The expression and transcriptional regulatory properties of the Autographa californica nuclear polyhedrosis virus IE1 gene product

Abstract

Gene regulation in the Autographa californica nuclear polyhedrosis virus is temporally regulated and sequentially ordered. Transcription is regulated, in part, by immediate early gene products with transcriptional regulatory properties. The IE1 gene product is a transcriptional regulator of AcMNPV genes. Studies were conducted to determine its expression throughout infection, and to elucidate novel transcriptional regulatory properties. The results of S1 nuclease assays showed that IE1 and IE0 RNAs were differentially expressed. Both mRNAs were expressed during the early phase of infection; whereas, only IE1 was expressed during the late phase. Two late spliced mRNAs containing the IE0 open reading frame were also identified. Novel activities of IE1 and IE0 were identified by transient cotransfection assays. The data showed that IE1 can function as an activator and repressor of gene expression. In addition to transactivating delayed early genes (Guarino and Summers, 1986a; 1986b), IE1 stimulated its own expression. The IE1 gene product also down-regulated expression of IE0. Although IE1 and IE0 are structurally similar, they expressed different activities. Unlike IE1, IE0 did not transactivate the delayed early gene 39K, without a cis-linked hr5 enhancer element. In addition, IE0 did not show any autoregulatory properties, but it did transactivate IE1. To further studies on the mechanism of IE1-mediated gene activation, a functional dissection of the IE1 gene product was conducted. Various mutants containing amino-terminal, carboxy-terminal, and internal deletions, and site specific mutations were constructed and tested for the ability to regulate gene expression and bind to the hr5 enhancer in vitro. The results indirectly showed that the IE1 gene product has two distinct and autonomous domains that regulate transcription and DNA binding. The amino terminal 145 amino acids contains a putative acidic activation domain required for transcriptional activation and repression, and the carboxy-terminal 100 amino acids contains a putative helix-loop-helix motif that mediates DNA binding activity. Mutants that expressed only the amino terminal acidic activation domain, and lacked the DNA binding domain, specifically interfered with wild type IE1 transactivation of the delayed early reporter plasmid 39CATQ- suggesting that this region interacts with transcriptional target proteins in vivo. The IE1 gene product functions as a transcriptional regulator of viral genes, and may be essential for the proper sequential activation and repression of genes throughout a viral infection.