Segmental copy number amplifications are more stable than aneuploidies in the absence of selection

Copy number variants (CNVs) are DNA duplications and deletions that cause genetic variation, underlying rapid adaptive evolution. CNVs often confer selective advantages, but can also incur fitness costs. Evolution of Saccharomyces cerevisiae in nutrient-limited chemostats recurrently selects for amplifications of nutrient transporter genes. However, their fate upon return to a non-selective environment remains unknown. To investigate CNV fitness and stability upon removing the original selection pressure, we studied 15 CNV lineages (11 segmental, 4 whole-chromosomal amplifications) selected in nitrogen-limited chemostats. CNV stability was monitored using fluorescent reporters during propagation in nutrient-rich batch cultures for 110–220 generations. All aneuploid lineages showed rapid CNV loss and reversion to a single-copy genotype, whereas segmental amplifications were remarkably stable– one of 11 strains reverted. Pairwise fitness competitions in rich media revealed strong fitness defects associated solely with CNVs that reverted; reversion led to increased fitness. Using simulation-based inference to estimate reversion rates and fitness effects, we determined negative selection as the primary driver of CNV loss. Whole-genome sequencing revealed that reversion of aneuploids and a segmental amplification left no evidence of prior CNV existence, rendering revertant genomes indistinguishable from the single-copy ancestor. Detailed characterization of a partial revertant identified chromosomal translocation, suggesting that extant CNVs can undergo structural diversification. Our findings provide novel evidence that most segmental CNVs adapted to nitrogen limitation are stable upon removal of selection, but costly gene amplifications are readily reversible. Together, these highlight the importance of CNVs in both long-term genome evolution and rapid, reversible adaptation to transient selection.