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Molecular and Cellular Biology, August 2000, p. 5847-5857, Vol. 20, No. 16
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Artificially Recruited TATA-Binding Protein Fails To Remodel Chromatin and Does Not Activate Three Promoters That Require Chromatin Remodeling

Michael P. Ryan,1 Grace A. Stafford,1 Liuning Yu,2 and Randall H. Morse1,2,*

Molecular Genetics Program, Wadsworth Center, New York State Department of Health,1 and SUNY School of Public Health,2 Albany, New York 12201-2002

Received 9 December 1999/Returned for modification 4 February 2000/Accepted 22 May 2000

Transcriptional activators are believed to work in part by recruiting general transcription factors, such as TATA-binding protein (TBP) and the RNA polymerase II holoenzyme. Activation domains also contribute to remodeling of chromatin in vivo. To determine whether these two activities represent distinct functions of activation domains, we have examined transcriptional activation and chromatin remodeling accompanying artificial recruitment of TBP in yeast (Saccharomyces cerevisiae). We measured transcription of reporter genes with defined chromatin structure by artificial recruitment of TBP and found that a reporter gene whose TATA element was relatively accessible could be activated by artificially recruited TBP, whereas two promoters, GAL10 and CHA1, that have accessible activator binding sites, but nucleosomal TATA elements, could not. A third reporter gene containing the HIS4 promoter could be activated by GAL4-TBP only when a RAP1 binding site was present, although RAP1 alone could not activate the reporter, suggesting that RAP1 was needed to open the chromatin structure to allow activation. Consistent with this interpretation, artificially recruited TBP was unable to perturb nucleosome positioning via a nucleosomal binding site, in contrast to a true activator such as GAL4, or to perturb the TATA-containing nucleosome at the CHA1 promoter. Finally, we show that activation of the GAL10 promoter by GAL4, which requires chromatin remodeling, can occur even in swi gcn5 yeast, implying that remodeling pathways independent of GCN5, the SWI-SNF complex, and TFIID can operate during transcriptional activation in vivo.

* Corresponding author. Mailing address: Molecular Genetics Program, Wadsworth Center, New York State Department of Health, Albany, NY 12201-2002. Phone: (518) 486-3116. Fax: (518) 474-3181. E-mail: Randall.Morse{at}wadsworth.org.

Molecular and Cellular Biology, August 2000, p. 5847-5857, Vol. 20, No. 16
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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