Separation of primary structural components conferring autoregulation, transactivation, and DNA-binding properties to the herpes simplex virus transcriptional regulatory protein ICP4.
A truncated ICP4 peptide which contains the amino-terminal 774 amino acids of the 1,298-amino-acid polypeptide is proficient for DNA binding, autoregulation, and transactivation of some viral genes (N. A. DeLuca and P. A. Schaffer, J. Virol. 62:732-743, 1988) and hence exhibits many of the properties characteristic of intact ICP4. To define the primary sequence important for the activities inherent in the amino-terminal half of the ICP4 molecule, insertional and deletion mutagenesis of the sequences encoding these residues were conducted. The DNA-binding activity of the molecule as assayed by the association with a consensus binding site was sensitive to insertional mutagenesis in two closely linked regions of the molecule. One region between amino acids 445 and 487 is critical for DNA binding and may contain a helix-turn-helix motif. The second region between amino acids 263 and 338 reduces the binding activity to a consensus binding site. When analyzed in the viral background, the DNA-binding activity of a peptide containing an insertion at amino acid 338 to a consensus binding site was reduced while the association with an alternative sequence was eliminated, suggesting a possible mechanism by which ICP4 may recognize a broader range of sequence elements. Mutations which eliminated DNA binding also eliminated or reduced both transactivation and autoregulation, supporting the requirement for DNA binding for these activities. Peptides that retained the deduced DNA-binding domain but lacked amino acids 143 through 210 retained the ability to associate with the consensus site and autoregulatory activity but were deficient for transactivation, demonstrating that the structural requirements for transactivation are greater than those required for autoregulation.
Laboratory of Tumor Virus Genetics, Dana-Farber Cancer Institute, Boston, Massachusetts.
A A
AA
Shepard
A N
AN
Imbalzano
N A
NA
DeLuca
D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000032:analysis*; D005786:Gene Expression Regulation; D006706:Homeostasis; D017874:Immediate-Early Proteins; D009154:Mutation; D011995:Recombination, Genetic; D014157:Transcription Factors / Q000032:analysis*; D014158:Transcription, Genetic; D014764:Viral Proteins / Q000032:analysis* / Q000235:genetics
D004268:DNA-Binding Proteins; D017874:Immediate-Early Proteins; D014157:Transcription Factors; D014764:Viral Proteins; C048925:herpes simplex virus, type 1 protein ICP4; D004247:DNA