Previous studies on a chromatin reporter gene (GAL-URARIB) in yeast showed that nucleosomes were maintained but rearranged during transcription in galactose, which was consistent with local dissociation of histones at the site of the RNA polymerase. Furthermore, repositioning of nucleosomes occurred rapidly after glucose repression. Because nucleosomal disruption and transcription produce topological changes in the chromatin substrate, the effect of topoisomerase activity was tested by the insertion of GAL-URABIB in topoisomerase mutant strains. The chromatin structure was analysed by nuclease digestion and psoralen crosslinking, and compared with that of the rDNA locus. In GAL-URARIB, neither the inactivation of topoisomerases I, II or I and II generated nucleosomal loss during transcription, nor was topoisomerase activity required for repositioning of the nucleosomes after repression. In contrast, the inactivation of topoisomerase I promoted an enhanced psoralen accessibility of the transcribed rDNA, possibly because of altered supercoiling, and the inactivation of topoisomerases I and II disrupted the chromatin structure of the whole rDNA locus by redistribution of the nucleosomes. The inactivation of topoisomerase II alone had no effect. These observations substantiate a differential participation of topoisomerases in the modulation of the chromatin structures of rDNA genes and of a single copy polymerase II gene. It is suggested that topological stress in genes transcribed by RNA polymerase II might diffuse away into flanking regions.

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Journal Article | Research Support, Non-U.S. Gov't

Authors

Cavalli G, Bachmann D, Thoma F

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