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J Virol. 1979 February; 29(2): 458-465

Autoregulation of Adenovirus Type 5 Early Gene Expression III. Transcription Studies in Isolated Nuclei

R. A. Blanton and T. H. Carter

¹ Department of Microbiology and Specialized Cancer Research Center, The Milton S. Hershey Medical Center, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania 17033

ABSTRACT

The rate of adenovirus RNA synthesis was compared in nuclei isolated from cells infected at 40.5°C in the presence of 1-ß-D-arabinofuranosylcytosine with adenovirus 5 or an early temperature-sensitive mutant of adenovirus type 5, H5ts125 (ts125). In nuclei isolated at various times after infection, the maximum amount of virus RNA synthesis occurred at 6 h after infection, after which time virus RNA synthesis declined in nuclei from wild-type infections but remained high in nuclei from ts125 infections. At 12 h after infection, the amount of virus RNA synthesis was 8- to 11-fold higher in nuclei from ts125 infections than in nuclei from wild-type infections. However, the kinetics of virus RNA synthesis in nuclei isolated from both infections were similar. When a ts125-infected culture was shifted to 32°C for 3 h (12 to 15 h after infection) before nucleus isolation, the amount of virus RNA synthesis in the isolated nuclei was reduced to nearly wild-type levels. A pulse-chase experiment showed little difference in degradation rates of virus RNA in isolated nuclei from wild-type and ts125 infections. Hybridization of RNA synthesized in vitro to restriction fragments of adenovirus type 5 DNA was consistent with early virus RNA. These results support the idea that the 72,000-dalton DNA-binding protein encoded by the mutant gene in ts125 can regulate early adenovirus gene expression by inhibiting initiation of transcription of the adenovirus genome.

Present address: Department of Biological Sciences, St. John's University, Jamaica, NY 11439.