THE SMALL-SUBUNIT OF THE VACCINIA VIRUS EARLY TRANSCRIPTION FACTOR CONTACTS THE TRANSCRIPTION PROMOTER DNA

The vaccinia virus early transcription factor (VETF), in addition to t he viral RNA polymerase, is required for efficient transcription of ea rly genes in vitro. VETF is a heterodimeric protein that binds specifi cally to early gene promoters. In order to localize the VETF DNA bindi ng domain, we have used photoreactive oligonucleotide probes with the sequence of the vaccinia virus growth factor promoter. The probes cons isted of double-stranded oligonucleotides incorporating radiolabeled d AMP and 5-bromo-dUMP into sequences of the promoter known to contact V ETF. Irradiation of a DNA probe having these nucleotides located upstr eam of the transcription start site in the presence of VFTF resulted i n the transfer of label to a polypeptide that comigrated with the smal l subunit of VETF. The label transfer reaction was shown to occur with the recombinant VETF small subunit in the absence of the large subuni t. These results indicate that the small subunit comprises at least pa rt of the VETF DNA binding domain and contacts the promoter in the reg ion upstream of the transcription start site.