Structural and functional characterization on the interaction of yeast TFIID subunit TAF1 with TATA-binding protein.
General transcription factor TFIID, consisting of TATA-binding protein (TBP) and TBP-associated factors (TAFs), plays a central role in both positive and negative regulation of transcription. The TAF N-terminal domain (TAND) of TAF1 has been shown to interact with TBP and to modulate the interaction of TBP with the TATA box, which is required for transcriptional initiation and activation of TATA-promoter operated genes. We have previously demonstrated that the Drosophila TAND region of TAF1 (residues 11-77) undergoes an induced folding from a largely unstructured state to a globular structure that occupies the DNA-binding surface of TBP thereby inhibiting the DNA-binding activity of TBP. In Saccharomyces cerevisiae, the TAND region of TAF1 displays marked differences in the primary structure relative to Drosophila TAF1 (11% identity) yet possesses transcriptional activity both in vivo and in vitro. Here we present structural and functional studies of yeast TAND1 and TAND2 regions (residues 10-37, and 46-71, respectively). Our NMR data show that, in yeast, TAND1 contains two alpha-helices (residues 16-23, 30-36) and TAND2 forms a mini beta-sheet structure (residues 53-56, 61-64). These TAND1 and TAND2 structured regions interact with the concave and convex sides of the saddle-like structure of TBP, respectively. Present NMR, mutagenesis and genetic data together elucidate that the minimal region (TAND1 core) required for GAL4-dependent transcriptional activation corresponds to the first helix region of TAND1, while the functional core region of TAND2, involved in direct interaction with TBP convex alpha-helix 2, overlaps with the mini beta-sheet region.
Mal
Tapas K
TK
Division of Molecular and Structural Biology, Ontario Cancer Institute and Department of Medical Biophysics, University of Toronto, Toronto, Ont. M5G 2M9, Canada.
Masutomi
Yutaka
Y
Zheng
Le
L
Nakata
Yasuto
Y
Ohta
Hiroshi
H
Nakatani
Yoshihiro
Y
Kokubo
Tetsuro
T
Ikura
Mitsuhiko
M
D000595:Amino Acid Sequence; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019906:Nuclear Magnetic Resonance, Biomolecular; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D017510:Protein Folding; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D035181:TATA-Box Binding Protein / Q000737:chemistry / Q000378:metabolism*; D035362:Transcription Factor TFIID / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
D035181:TATA-Box Binding Protein; D035362:Transcription Factor TFIID