Transcriptional Activity and Chromatin Structure of Enhancer-Deleted rRNA Genes in Saccharomyces cerevisiae

Abstract

We used the psoralen gel retardation assay and Northern blot analysis in an in vivo yeast system to analyze effects of rDNA enhancer deletions on the chromatin structure and the transcription of tagged rDNA units. We found that upon deletion of a single enhancer element, transcription of the upstream and downstream rRNA gene was reduced by about 50%. Although removing both flanking enhancers of an rRNA gene led to a further reduction in transcription levels, a significant amount of transcriptional activity remained, either resulting from the influence of more distantly located enhancer elements or reflecting the basal activity of the polymerase I promoter within the nucleolus. Despite the reduction of transcriptional activity upon enhancer deletion, the activation frequency (proportion of nonnucleosomal to nucleosomal gene copies in a given cell culture) of the tagged rRNA genes was not significantly altered, as determined by the psoralen gel retardation assay. This is a strong indication that, within the nucleolus, the yeast rDNA enhancer functions by increasing transcription rates of active rRNA genes and not by activating silent transcription units.

ACKNOWLEDGMENTS

We thank F. Thoma for correction of the manuscript, C. Weissmann for access to the PhosphorImager; R. Wellinger, B. Schweizer, V. Taylor, and M. Smerdon for helpful discussion; and U. Suter for continuous support. We also thank the anonymous referees for helpful suggestions.

This work was supported by grants from the Boehringer Ingelheim Fonds (to M.B.) and the Swiss National Science Foundation (to J.M.S.; grant 31-52246.97).