DNA-binding and transcriptional properties of human transcription factor TFIID after mild proteolysis.
The existence of separable functions within the human class II general transcription factor TFIID was probed for differential sensitivity to mild proteolytic treatment. Independent of whether TFIID was bound to DNA or free in solution, partial digestion with either one of a variety of nonspecific endoproteases generated a protease-resistant protein product that retained specific DNA recognition, as revealed by DNase I footprinting. However, in contrast to native TFIID, which interacts with the adenovirus major late (ML) promoter over a very broad DNA region, partially proteolyzed TFIID interacted with only a small region of the ML promoter immediately surrounding the TATA sequence. This novel footprint was very similar to that observed with the TATA factor purified from yeast cells. Partially proteolyzed human TFIID could form stable complexes that were resistant to challenge by exogenous templates. It could also nucleate the assembly of transcription complexes on the ML promoter with an efficiency comparable to that of native TFIID, yielding similar levels of transcription initiation. These results suggest a model in which the human TFIID protein is composed of at least two different regions or polypeptides: a protease-resistant "core," which by itself is sufficient for promoter recognition and basal transcriptional levels, and a protease-sensitive "tail," which interacts with downstream promoter regions and may be involved in regulatory processes.
Department of Tumor Biology, University of Texas M. D. Anderson Cancer Center, Houston 77030.
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D000256:Adenoviridae / Q000235:genetics; D002467:Cell Nucleus / Q000378:metabolism; D004279:DNA, Viral / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003850:Deoxyribonuclease I; D006367:HeLa Cells / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D010446:Peptide Fragments / Q000378:metabolism; D010447:Peptide Hydrolases / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D035362:Transcription Factor TFIID; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic
D004279:DNA, Viral; D004268:DNA-Binding Proteins; D010446:Peptide Fragments; D035362:Transcription Factor TFIID; D014157:Transcription Factors; D003850:Deoxyribonuclease I; D010447:Peptide Hydrolases