Queitsch, ChristineHall, Ashley Nicole2022-01-262022-01-262021Hall_washington_0250E_23675.pdfhttp://hdl.handle.net/1773/48300Thesis (Ph.D.)--University of Washington, 2021The ribosomal DNA (rDNA) encodes the ribosomal RNAs and is present in tens to thousands of tandemly repeated copies in eukaryotic genomes. rDNA copy number varies among individual humans and within model organisms. Having too few rDNA copies restricts ribosome biogenesis and leads to reduced growth rate or death. Variation within the naturally occurring range of rDNA copy numbers likely leaves ribosome biogenesis intact but still confers cellular phenotypes, such as altering the global transcriptome in humans. The extent to which natural rDNA copy number variation impacts whole-organism phenotypes such as fitness and lifespan is poorly understood.A prerequisite to understanding the impact of rDNA copy number on phenotype is accurate copy number measurement. The gold standard for rDNA copy number measurement is pulsed-field gel electrophoresis and Southern blotting. Because this method is low-throughput and ill-suited for human rDNA measurement, many studies rely on short-read sequencing data. However, we found that sequencing-based rDNA copy number estimates are highly error-prone. This high technical error rate directly impacts conclusions: A study assessing 168 low coverage sequencing samples concluded that the 5S and 45S rDNA arrays co-vary in copy number in humans. With thousands of samples from newer, higher quality datasets, I demonstrated that there is no meaningful co-variation between the 5S and 45S array copy numbers in humans. I further generated resources of Caenorhabditis elegans with rDNA copy numbers from the high and the low end of the worm’s natural range, including a set of recombinant inbred lines. Changing rDNA copy number did not result in a detectable change in rRNA abundance, consistent with these copy numbers supporting functional ribosome biogenesis. I found that in the naturally occurring rDNA copy number range of C. elegans, copy number differences confer no aging or fitness defects. In addition, this rDNA copy number variation confers few differences in global gene expression. These results suggest that any phenotypic consequences of rDNA copy number variation in the naturally occurring range are subtle, at least in C. elegans, and to achieve substantial phenotypic consequence, rDNA copy number must be pushed to extremely high or low levels.application/pdfen-USCC BYCopy number variationLifespanrDNARibosomal DNARibosomeGeneticsMolecular and cellular biologyThesis