The Highly Conserved Basic Domain I of Baculovirus IE1 Is Required for hr Enhancer DNA Binding and hr -Dependent Transactivation
Open Access
- 15 May 2003
- journal article
- Published by American Society for Microbiology in Journal of Virology
- Vol. 77 (10), 5668-5677
- https://doi.org/10.1128/jvi.77.10.5668-5677.2003
Abstract
The immediate-early protein IE1 is the principal transcriptional regulator of the baculovirus Autographa californica nucleopolyhedrovirus (Ac M NPV). Transactivation by IE1 is dramatically stimulated by cis linkage of the affected promoter to Ac M NPV homologous region ( hr ) elements that contain palindromic 28-bp repeats (28-mers) with enhancer activity. This hr -dependent transcriptional enhancement requires binding of the 28-mer by dimeric IE1. Here, we have defined IE1 domains required for this DNA binding in order to investigate the mechanism of IE1 function. Analysis of a panel of IE1 insertion mutations indicated that disruption of a highly conserved domain (residues 152 to 161) consisting of mostly positive-charged residues (basic domain I) abolished hr -dependent transactivation. Targeted mutagenesis of basic residues within basic domain I caused loss of hr -dependent transactivation but had no effect on IE1 oligomerization, nuclear localization, or hr -independent transactivation of viral promoters. Alanine substitutions of K 152 and K 154 or K 160 and K 161 impaired IE1 binding to 28-mer DNA as a homodimer, indicating that these basic residues are required for enhancer binding. Consistent with a DNA-binding defect, 28-mer interaction was improved by heterodimerization with wild-type IE1 or by increasing mutated IE1 concentrations. DNA binding mediated by basic domain I was also required for IE1 transactivation that occurred through physically separated, unlinked hr elements. We concluded that basic domain I is the enhancer-binding domain for IE1. Our data also suggest that DNA binding activates IE1 for transcriptional enhancement, possibly through a conformational change involving basic domain I.Keywords
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