title: Characterization of the acidic domain of the IE1 regulatory protein from Orgyia pseudotsugata multicapsid nucleopolyhedrovirus.
abstract: This study presents a detailed analysis of the acidic N-terminal region of the Orgyia pseudotsugata multicapsid nucleopolyhedrovirus (OpMNPV) transactivator IE1. The N-terminal region of IE1 is rich in acidic amino acids and has been hypothesized to be an acidic activation domain. Removal of the N-terminal 126 amino acids containing the acidic domain of IE1 resulted in complete loss of transactivation activity, indicating that this region is essential for transactivation. The OpMNPV acidic domain was replaced with the archetype acidic activation domain from VP16 and the acid-rich region of Autographa californica multicapsid NPV (AcMNPV) IE1. These chimeric constructs were fully capable of transactivation in transient assays. The chimeric OpMNPV IE1s containing the herpes simplex virus VP16 and AcMNPV IE1 acidic activation domains consistently transactivated a reporter gene to higher levels than the OpMNPV IE1 acidic activation domain. Transactivation by the chimeric constructs is enhanced synergistically when cotransfected with IE2 into Lymantria dispar and Spodoptera frugiperda cells. Both N- to C-terminal and C- to N-terminal deletions of the OpMNPV acidic activation domain were constructed to define functional domains within the OpMNPV IE1 acidic activation domain. At least two potential activation domains were identified. Within each of these domains, two core regions at amino acids 28-43 and amino acids 113-124 were identified that were similar to core regions of VP16 and GAL4, which contain predominately acidic and bulky hydrophobic amino acids.
authors.affiliation: Department of Zoology, University of British Columbia, Vancouver, Canada.
authors.forename: I J
authors.initials: IJ
authors.lastname: Forsythe
authors.forename: C E
authors.initials: CE
authors.lastname: Shippam
authors.forename: L G
authors.initials: LG
authors.lastname: Willis
authors.forename: S
authors.initials: S
authors.lastname: Stewart
authors.forename: T
authors.initials: T
authors.lastname: Grigliatti
authors.forename: D A
authors.initials: DA
authors.lastname: Theilmann
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics; D017405:Herpes Simplex Virus Protein Vmw65 / Q000737:chemistry / Q000235:genetics; D017874:Immediate-Early Proteins / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D017924:Nucleopolyhedroviruses / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D016415:Sequence Alignment; D018411:Spodoptera; D015534:Trans-Activators / Q000737:chemistry* / Q000235:genetics
chemical_list: C069183:Ac-IE-1 protein, Autographa californica nucleopolyhedrovirus; D004268:DNA-Binding Proteins; D017405:Herpes Simplex Virus Protein Vmw65; D017874:Immediate-Early Proteins; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; C072773:IE2 protein, polyhedrosis virus