DNA replication stress restricts ribosomal DNA copy number

• Published in: PLoS genetics Vol. 13; no. 9; p. e1007006
• Main Authors: Salim, Devika, Bradford, William D, Freeland, Amy, Cady, Gillian, Wang, Jianmin, Pruitt, Steven C, Gerton, Jennifer L
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 15-09-2017; Public Library of Science (PLoS)
• Subjects: Animals; Baking yeast; Bioinformatics; Biology and life sciences; Biosynthesis; Cancer; Cell Cycle Proteins - genetics; Cellular biology; Chromosomes; Contraction; Copy number; Deoxyribonucleic acid; DNA; DNA biosynthesis; DNA Copy Number Variations - genetics; DNA replication; DNA Replication - genetics; DNA, Ribosomal - genetics; DNA-Binding Proteins - genetics; Funding; Genetic aspects; Genomes; Medical research; Mice; Minichromosome Maintenance Complex Component 2 - genetics; Mutation; Nucleotide sequence; Observations; Physical Sciences; Recombination, Genetic; Replication; Research and analysis methods; Ribosomal DNA; RNA polymerase; Saccharomyces cerevisiae; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae Proteins - genetics; Stem cells; Temperature; Temperature-sensitive mutant; Tumors; Yeast; Yeasts (Fungi); United States > US; Kansas City Missouri
• ISSN: 1553-7404; 1553-7390; 1553-7404
• DOI: 10.1371/journal.pgen.1007006

Ribosomal RNAs (rRNAs) in budding yeast are encoded by ~100-200 repeats of a 9.1kb sequence arranged in tandem on chromosome XII, the ribosomal DNA (rDNA) locus. Copy number of rDNA repeat units in eukaryotic cells is maintained far in excess of the requirement for ribosome biogenesis. Despite the importance of the repeats for both ribosomal and non-ribosomal functions, it is currently not known how "normal" copy number is determined or maintained. To identify essential genes involved in the maintenance of rDNA copy number, we developed a droplet digital PCR based assay to measure rDNA copy number in yeast and used it to screen a yeast conditional temperature-sensitive mutant collection of essential genes. Our screen revealed that low rDNA copy number is associated with compromised DNA replication. Further, subculturing yeast under two separate conditions of DNA replication stress selected for a contraction of the rDNA array independent of the replication fork blocking protein, Fob1. Interestingly, cells with a contracted array grew better than their counterparts with normal copy number under conditions of DNA replication stress. Our data indicate that DNA replication stresses select for a smaller rDNA array. We speculate that this liberates scarce replication factors for use by the rest of the genome, which in turn helps cells complete DNA replication and continue to propagate. Interestingly, tumors from mini chromosome maintenance 2 (MCM2)-deficient mice also show a loss of rDNA repeats. Our data suggest that a reduction in rDNA copy number may indicate a history of DNA replication stress, and that rDNA array size could serve as a diagnostic marker for replication stress. Taken together, these data begin to suggest the selective pressures that combine to yield a "normal" rDNA copy number.