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Evidence that TAF-TATA Box-Binding Protein Interactions Are Required for Activated Transcription in Mammalian Cells

Department of Microbiology, Immunology, and Molecular Genetics,¹ Department of Molecular Pharmacology,² Molecular Biology Institute, University of California, Los Angeles, Los Angeles, California³

Received 27 August 2001/ Returned for modification 8 October 2001/ Accepted 20 December 2001

Surfaces of human TATA box-binding protein (hsTBP) required for activated transcription in vivo were defined by constructing a library of surface residue substitution mutations and assaying them for their ability to support activated transcription in transient-transfection assays. In earlier work, three regions were identified where mutations inhibited activated transcription without interfering with TATA box DNA binding. One region is on the upstream surface of the N-terminal TBP repeat with respect to the direction of transcription and corresponds to the TBP surface that interacts with TFIIA. A second region on the stirrup of the C-terminal TBP repeat corresponds to the TFIIB-binding surface. Here we report that the third region where mutations inhibit activated transcription in mammalian cells, the convex surface of the N-terminal repeat, corresponds to a surface on TBP that interacts with hsTAF1, the major scaffold subunit of TFIID. Since mutations at the center of the hsTAF1-interacting region inhibit the ability of the protein to support activated transcription in vivo, these results are consistent with the conclusion that an interaction between hsTBP and TAF_IIs is required for activated transcription in mammalian cells.

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