TFIIA is a transcription factor that, by interacting with the TATA-binding subunit (TBP) of TFIID, modulates transcription initiation by RNA polymerase II in vitro. By use of a mobility shift assay, TFIIA was purified from HeLa cells as a complex of 35-, 19-, and 12-kD subunits. Oligonucleotides were used to isolate a human cDNA clone, hTFIIA/alpha, which encodes a 55-kD protein with homology to the product of the yeast gene TOA1. The open reading frame of hTFIIA/alpha contains peptide sequences obtained from both the p35 and p19 subunits of natural human TFIIA, and thus encodes these two subunits. Consistent with this, antiserum raised against the 55-kD hTFIIA/alpha-encoded protein reacted with both the p35 and p19 subunits of natural TFIIA, and the recombinant protein could functionally replace those subunits in a mobility shift assay with renatured p12. An efficient affinity purification for natural human TFIIA was suggested by the sequence of the hTFIIA/alpha protein and demonstrated biochemically. Finally, transcription from the adenovirus major late promoter was greatly reduced in nuclear extracts depleted with anti-TFIIA/alpha serum and was restored to original levels by the readdition of purified human TFIIA.

• PMID: 8224848
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Reference Type

Comparative Study | Journal Article | Research Support, Non-U.S. Gov't | Research Support, U.S. Gov't, P.H.S.

Authors

DeJong J, Roeder RG

Primary Lit For

Transcription factor TFIIA complex | General transcription factor complex TFIID | DNA-directed RNA polymerase II complex

Additional Lit For

TOA1

Review For