The CREB constitutive activation domain interacts with TATA-binding protein-associated factor 110 (TAF110) through specific hydrophobic residues in one of the three subdomains required for both activation and TAF110 binding.
The cAMP response element-binding protein (CREB) mediates both basal and PKA-inducible transcription through two separate and independently active domains, the constitutive activation domain (CAD) and the kinase-inducible domain, respectively. The CREB CAD interacts with the general transcription factor TFIID through one or more of the TATA-binding protein-associated factors (TAFs), one of which is TAF110. The CAD is composed of three subdomains, rich in either serine, hydrophobic amino acids, or glutamine. In the present study, analysis of deletion mutants of the CAD showed that all three CAD subdomains were required for effective interaction with TAF110 in a yeast two-hybrid assay. Therefore, a library of random point mutations within the CAD was analyzed in a reverse two-hybrid screen to identify amino acids that are essential for interaction with the TAF. Interaction defects resulted solely from mutations of hydrophobic amino acid residues within the hydrophobic cluster to charged amino acid residues. Together, the deletion and mutation analyses suggest that the entire CAD provides an environment for a specific hydrophobic interaction with TAF110 that is crucial for interaction. Our results provide further evidence for a model of basal activation by CREB involving interaction with TAF110 that promotes recruitment or stabilization of TFIID binding to the promoter, which facilitates pre-initiation complex assembly.
Felinski
E A
EA
Department of Cellular and Molecular Physiology and the Cell and Molecular Biology Program, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania 17033, USA.
Quinn
P G
PG
D017362:Cyclic AMP Response Element-Binding Protein / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D029721:Drosophila Proteins; D016296:Mutagenesis; D017354:Point Mutation; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000235:genetics; D035381:TATA-Binding Protein Associated Factors; D035181:TATA-Box Binding Protein; D035362:Transcription Factor TFIID; D014157:Transcription Factors / Q000378:metabolism*
D017362:Cyclic AMP Response Element-Binding Protein; D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D035381:TATA-Binding Protein Associated Factors; D035181:TATA-Box Binding Protein; C103338:Taf4 protein, Drosophila; D035362:Transcription Factor TFIID; D014157:Transcription Factors