PgmNr Y3075: Unsolicited rDNA copy number variants frequently occur in yeast deletion collections and transformations.

Authors:
Elizabeth X. Kwan 1 ; Xiaobin S. Wang 1,2 ; Haley M. Amemiya 1 ; M. K. Raghuraman 1 ; Bonita J. Brewer 1


Institutes
1) University of Washington, Seattle, WA; 2) Columbia University, New York, NY.


Keyword: 02. Chromosome structure, variation, evolution and dynamics

Abstract:

Ribosomal DNA (rDNA) is present in many copies in eukaryotic organisms and has been recently implicated in the regulation of gene expression, responses to replication stress, telomeric silencing, and aging.  The budding yeast S. cerevisiae maintains 150 rDNA copies in a tandem array on chromosome XII.  Instability of the yeast rDNA array is well documented, but the rDNA locus typically returns to wildtype copy numbers after perturbation, suggesting internal regulation of copy number maintenance.  We sought to identify determinants of yeast rDNA copy number by performing a screen using the Yeast Knock-Out (YKO) single gene deletion collection.  We also wanted to assess the relationship between rDNA number and yeast replicative lifespan and therefore examined the rDNA sizes of 434 YKO strains, 221 of which were reported to have extended replicative lifespans.  We found no correlation between yeast longevity and rDNA copy number, but did identify 64 YKO candidates with significantly changed rDNA copy numbers.  However, in the process of validating candidates using de novo gene deletions and tetrad analysis, we found that the altered rDNA phenotypes in the candidate YKO strains were not associated with the deleted genes.  Instead, we found that rDNA copy number variants (CNVs) appeared at random within multiple transformants from each de novo gene deletion.  Further examination revealed that cells taken through standard transformation protocols generated rDNA CNVs and that exposure to lithium acetate alone increased the rate of observed rDNA CNVs.  Since lithium acetate transformation is commonly used in yeast strain construction, unanticipated rDNA CNVs could be potential passenger mutations that may have repercussions in the increasing number of reported cellular processes in which rDNA copy number matters.