Originally published as Genetics Published Articles Ahead of Print on March 6, 2009.

Genetics, Vol. 182, 105-119, May 2009, Copyright © 2009
doi:10.1534/genetics.108.100313

Genetic Identification of Factors That Modulate Ribosomal DNA Transcription in Saccharomyces cerevisiae

Robert D. Hontz, Rachel O. Niederer, Joseph M. Johnson and Jeffrey S. Smith1

Department of Biochemistry and Molecular Genetics, University of Virginia Health System, School of Medicine, Charlottesville, Virginia 22908

1 Corresponding author: Department of Biochemistry and Molecular Genetics, University of Virginia Health System, School of Medicine, Jordan Hall, Box 800733, Charlottesville, VA 22908.
E-mail: jss5y{at}virginia.edu

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).