Defects in the mitochondrial respiratory chain (RC) underlie a spectrum of human conditions, ranging from devastating inborn errors of metabolism to aging. We performed a genome-wide, Cas9-mediated screen to identify factors that are protective during RC inhibition. Our results highlight the hypoxia response, an endogenous program evolved to adapt to limiting oxygen availability. Genetic or small molecule activation of the hypoxia response is protective against mitochondrial toxicity in cultured cells and zebrafish models. Chronic hypoxia leads to a marked improvement in survival, body weight, body temperature, behavior, neuropathology and disease biomarkers in a genetic mouse model of Leigh syndrome, the most common pediatric manifestation of mitochondrial disease. Further preclinical studies are required to assess whether hypoxic exposure can be developed into a safe and effective treatment for human diseases associated with mitochondrial dysfunction.