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Molecular and Cellular Biology, April 1999, p. 2817-2827, Vol. 19, No. 4
0270-7306/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

An In Vitro System Recapitulates Chromatin Remodeling at the PHO5 Promoter

Elizabeth S. Haswell and Erin K. O'Shea*

Department of Biochemistry and Biophysics, University of California, San Francisco, School of Medicine, San Francisco, California 94143-0448

Received 14 December 1998/Accepted 16 December 1998

The Saccharomyces cerevisiae gene PHO5 is an excellent system with which to study regulated changes in chromatin structure. The PHO5 promoter is packaged into four positioned nucleosomes under repressing conditions; upon induction, the structure of these nucleosomes is altered such that the promoter DNA becomes accessible to nucleases. We report here the development and characterization of an in vitro system in which partially purified PHO5 minichromosomes undergo promoter chromatin remodeling. Several hallmarks of the PHO5 chromatin transition in vivo were reproduced in this system. Chromatin remodeling of PHO5 minichromosomes required the transcription factors Pho4 and Pho2, was localized to the promoter region of PHO5, and was independent of the chromatin-remodeling complex Swi-Snf. In vitro chromatin remodeling also required the addition of fractionated nuclear extract and hydrolyzable ATP. This in vitro system should serve as a useful tool for identifying the components required for this reaction and for elucidating the mechanism by which the PHO5 promoter chromatin structure is changed.

* Corresponding author. Mailing address: Dept. of Biochemistry and Biophysics, Box 0448, University of California, San Francisco, School of Medicine, San Francisco, CA 94143-0448. Phone: (415) 476-2212. Fax: (415) 502-4315. E-mail: oshea{at}biochem.ucsf.edu.

Molecular and Cellular Biology, April 1999, p. 2817-2827, Vol. 19, No. 4
0270-7306/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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