Genetic identification of factors that modulate ribosomal DNA transcription in Saccharomyces cerevisiae.
Ribosomal RNA (rRNA) is transcribed from the ribosomal DNA (rDNA) genes by RNA polymerase I (Pol I). Despite being responsible for the majority of transcription in growing cells, Pol I regulation is poorly understood compared to Pol II. To gain new insights into rDNA transcriptional regulation, we developed a genetic assay in Saccharomyces cerevisiae that detects alterations in transcription from the centromere-proximal rDNA gene of the tandem array. Changes in Pol I transcription at this gene alter the expression of an adjacent, modified URA3 reporter cassette (mURA3) such that reductions in Pol I transcription induce growth on synthetic media lacking uracil. Increases in Pol I transcription induce growth on media containing 5-FOA. A transposon mutagenesis screen was performed with the reporter strain to identify genes that play a role in modulating rDNA transcription. Mutations in 68 different genes were identified, several of which were already known to function in chromatin modification and the regulation of Pol II transcription. Among the other classes of genes were those encoding proteasome subunits and multiple kinases and phosphatases that function in nutrient and stress signaling pathways. Fourteen genes were previously uncharacterized and have been named as regulators of rDNA transcription (RRT).
Hontz
Robert D
RD
Department of Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, Virginia 22908, USA.
Niederer
Rachel O
RO
Johnson
Joseph M
JM
Smith
Jeffrey S
JS
D015153:Blotting, Western; D002843:Chromatin / Q000235:genetics*; D047369:Chromatin Immunoprecipitation; D004251:DNA Transposable Elements / Q000235:genetics; D004275:DNA, Ribosomal / Q000235:genetics*; D006655:Histone Deacetylases / Q000378:metabolism; D016296:Mutagenesis; D012318:RNA Polymerase I / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic
D002843:Chromatin; D004251:DNA Transposable Elements; D004275:DNA, Ribosomal; D012333:RNA, Messenger; D012318:RNA Polymerase I; D006655:Histone Deacetylases