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Volume 270, Number 11, Issue of March 17, 1995 pp. 6205-6210
©1995 by The American Society for Biochemistry and Molecular Biology, Inc.
Characterization of the Components of Reconstituted Saccharomyces cerevisiae RNA Polymerase I Transcription Complexes

(Received for publication, November 8, 1994; and in revised form, January 11, 1995)

Daniel L. Riggs ,  Cheryl L. Peterson,  J. Quyen Wickham,  Letrisa M. Miller,  Eileen M. Clarke,  John A. Crowell ,  Jean-Christophe Sergere

We have reconstituted specific RNA polymerase I transcription from three partially purified chromatographic fractions (termed A, B, and C). Here, we present the chromatographic scheme and the initial biochemical characterization of these fractions. The A fraction contained the RNA polymerase I transcription factor(s), which was necessary and sufficient to form stable preinitiation complexes at the promoter. Of the three fractions, only fraction A contained a significant amount of the TATA binding factor. The B fraction contributed RNA polymerase I, and it contained an essential RNA polymerase I transcription factor that was specifically inactivated in response to a significant decrease in growth rate. The function of the C fraction remains unclear. This reconstituted transcription system provides a starting point for the biochemical dissection of the yeast RNA polymerase I transcription complex, thus allowing in vitro experiments designed to elucidate the molecular mechanisms controlling rRNA synthesis.



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