The purpose of this study is to characterize the physiological and biochemical properties of the viral polypeptide designated ICP4 which is synthesized soon after infection of human cells by herpes simplex virus. HSV is the causative agent in humans of cold sores (herpes labialis) and genital lesions (herpes genitalis), both of which occur as a consequence of reactivation of a latent virus. HSV has been associated with carcinoma of the cervix. Previous work by others suggest that ICP4 functions as a regulatory protein to control the expression of several viral genes necessary for virus production in infected cells. In this investigation, two approaches will be used to elucidate the molecular processes which act to control and coordinate gene expression in HSV infected cells. The first approach will involve a study of the sequence of synthetic events which occur in cells infected with temperature-sensitive mutants of HSV that produce functionally impaired forms of ICP4 at the nonpermissive temperature. Induction of the transcription of beta polypeptide genes will be assayed by the Southern blot hybridization method using 32P-labeled RNA and HSV DNA restriction endonuclease fragments. Induction of the synthesis of beta polypeptides will be assayed enzymatically and by SDS-polyacrylamide gel electrophoresis. These results will be compared with those obtained using HSV infected cells treated with drugs which specifically inhibit RNA or protein synthesis. The second approach will involve the biochemical and biophysical characterization of ICP4 purified from extracts of HSF infected cells. The partial proteolytic digestion products of the three intracellular forms of ICP4 will be compared. ICP4 is a phosphoprotein; thus the structure of the bound phosphate moiety will be determined. ICP4 is a nuclear protein; thus the association of ICP4 with nuclear components such as chromatin and RNA will be analyzed. Using purified preparations of ICP4, the number and location of DNA-binding sites on the viral genome will be determined. The nucleotide sequence of these ICP4 binding sites will be obtained using the DNAase footprinting technique. The results gained from these studies will be used to construct a model for the regulation by ICP4 of beta gene expression in HSV-infected cells.