The
rDNA cluster is the genetic locus encoding the ribosomal RNAs and physically defines where ribosomes begin to be assembled. In the yeast Saccharomyces cerevisiae, the highly repetitive structure of this locus makes it a very interesting target for studies about
genome stability,
chromatin-mediated transcriptional silencing and progression of aging. In fact, recombination among the repeated units is suppressed in a WT cell. Moreover, when genes transcribed by
RNA polymerase II are inserted in the
rDNA cluster, their transcription is silenced. Finally, the formation of
rDNA minicircles (ERCs) has been shown to be one of the causes of aging in yeast.
DNA topoisomerase I have been shown to suppress recombination specifically at the
rDNA of S.cerevisiae. Moreover, also the
chromatin structure of this locus is affected in a top1 strain, because
rDNA specific transcriptional silencing is abolished. Nonetheless, the molecular basis of how this
enzyme interferes with these functions is yet unknown. Here are reported results obtained by in vivo studies of
DNA protein interactions occurring on the
rDNA locus. The analyses include a fine mapping of
nucleosome positioning;
RNA polymerase I transcription factors and
DNA topoisomerase I cleavage sites. Important conclusions can be drawn: i)
nucleosome positioning in the Non Transcribed Spacer is not affected by
RNA polymerase I transcription; ii) the
RNA polymerase I transcription factors bind
DNA in vivo with a defined hierarchy; iii) the
DNA topoisomerase I cleaves the NTS in very specific sites, but cleavage is not induced by
RNA polymerase I transcription. These in vivo studies help to characterize the molecular basis of important phenomena as the transcriptional silencing and
genome stability in yeast.