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Enhancement of TBP binding by activators and general transcription factors

Nature volume 399, pages 605–609 (1999)Cite this article

Abstract

Eukaryotic transcriptional activators function, at least in part, by promoting assembly of the preinitiation complex1,2,3, which comprises RNA polymerase II and its general transcription factors (GTFs). Activator-mediated stimulation of the assembly of the preinitiation complex has been studied in vitro but has been relatively refractory to in vivo analysis. Here we use a DNA-crosslinking/immunoprecipitation assay to study in living cells the first step in the assembly of the preinitiation complex, the interaction between the TATA-box-binding protein (TBP) and its binding site, the TATA box. Analysis of a variety of endogenous yeast genes, and of a series of activators of differing strength, reveals a general correlation between TBP binding and transcriptional activity. Using mutant yeast strains, we show that Mot1 prevents the binding of TBP to inactive promoters and that activator-mediated stimulation of TBP binding requires additional GTFs, including TFIIB and Srb4. Taken together, our results indicate that TBP binding in vivo is stringently controlled, and that the ability of activators to stimulate this step in the assembly of the preinitiation complex is a highly cooperative process involving multiple transcription factors.

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Figure 1: An in vivo crosslinking assay for measuring TBP–TATA-box interaction.
Figure 2: Correlation between TBP binding and transcriptional activit in vivo.
Figure 3: Correlation between transcriptional activity and association of TFIIB and RNA polymerase II with promoters.
Figure 4: Mot1 prevents TBP binding to inactive promoters.
Figure 5: Transcription components that facilitate activator-mediated stimulation of TBP binding.

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Acknowledgements

We thank R. Young, M. Grunstein, J. Thorner and J. Majors for yeast strains, J. Ma for plasmids, and K. Struhl for discussion and for communicating unpublished results. This work was supported in part by a grant from the NIH to M.R.G. M.R.G. is an investigator of the Howard Hughes Medical Institute.

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  1. Howard Hughes Medical Institute, Program in Molecular Medicine, University of Massachusetts Medical Center, 373 Plantation Street, Suite 309, Worcester, 01605, Massachusetts, USA

    Xiao-Yong Li, Amy Virbasius, Xiaochuan Zhu & Michael R. Green

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  1. Xiao-Yong Li
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  2. Amy Virbasius
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Correspondence to Michael R. Green.

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Li, XY., Virbasius, A., Zhu, X. et al. Enhancement of TBP binding by activators and general transcription factors. Nature 399, 605–609 (1999). https://doi.org/10.1038/21232

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