A single cDNA, hTFIIA/alpha, encodes both the p35 and p19 subunits of human TFIIA.
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.
DeJong
J
J
Laboratory of Biochemistry and Molecular Biology, Rockefeller University, New York, New York 10021.
Roeder
R G
RG
D000595:Amino Acid Sequence; D001483:Base Sequence; D002467:Cell Nucleus / Q000378:metabolism; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D017124:Conserved Sequence; D018076:DNA, Complementary / Q000378:metabolism*; D015870:Gene Expression; D006367:HeLa Cells; D006801:Humans; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D035561:Transcription Factor TFIIA; D035362:Transcription Factor TFIID; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D014158:Transcription, Genetic
D018076:DNA, Complementary; D046911:Macromolecular Substances; D009838:Oligodeoxyribonucleotides; D029701:Saccharomyces cerevisiae Proteins; C441519:TOA1 protein, S cerevisiae; D035561:Transcription Factor TFIIA; D035362:Transcription Factor TFIID; D014157:Transcription Factors