show Abstracthide AbstractAneuploidy, in which cells carry an abnormal chromosome count, is detrimental during development yet common in human cancers; why cells differ in tolerance remains unclear. We mapped the genetic basis of aneuploidy tolerance in wild Saccharomyces cerevisiae versus the sensitive lab strain to Ssd1, an RNA-binding protein involved in translation whose loss recapitulates aneuploidy signatures in laboratory yeast. We find Ssd1 localizes to mitochondria, influences localization of nuclear-encoded mitochondrial mRNAs and/or abundance of the encoded proteins, influences mitochondrial function, and minimizes protein aggregates upon chromosome amplification. Recapitulating ssd1D defects with combinatorial drug treatment selectively targets wild-type aneuploids in multiple strains, suggesting therapeutic approaches. Our work adds to elegant studies done in the sensitized laboratory strain to present a mechanistic understanding of aneuploidy tolerance in eukaryotes. Overall design: RNA-seq and transcriptome analysis of S. cerevisiae aneuploids YPS1009, NCYC110 having SSD1 deleted. Biological triplicate was minimally performed for RNA-seq. RIP-seq analysis of YPS1009 having SSD1 tagged with GFP, in biological duplicate.