The γ‐secretase complex (which contains presenilins, nicastrin, anterior pharynx defective‐1, and presenilin enhancer‐2) cleaves type I transmembrane proteins, including Notch and amyloid precursor protein. Dysregulated γ‐secretase activity has been implicated in the pathogenesis of Alzheimer's disease, stroke, atherosclerosis, and cancer. Tight regulation of γ‐secretase activity is required for normal physiology. Here, we isolated HIG1 (hypoxia inducible gene 1, domain member 1A) from a functional screen of γ‐secretase inhibitory genes. HIG1 was highly expressed in the brain. Interestingly, HIG1 was localized to the mitochondria and was directly bound to γ‐secretase components on the mitochondrial membrane in SK‐N‐SH neuroblastoma cells. Overexpresssion of HIG1 attenuated hypoxia‐induced γ‐secretase activation on the mitochondrial membrane and the accumulation of intracellular amyloid β. This accumulation was accompanied by hypoxia‐induced mitochondrial dysfunction. The latter half domain of HIG1 was required for binding to the γ‐secretase complex and suppression of γ‐secretase activity. Moreover, depletion of HIG1 increased γ‐secretase activation and enhanced hypoxia‐induced mitochondrial dysfunction. In summary, HIG1 is a novel modulator of the mitochondrial γ‐secretase complex, and may play a role in the maintenance of normal mitochondrial function.—Hayashi, H., Nakagami, H., Takeichi, M., Shimamura, M., Koibuchi, N., Oiki, E., Sato, N., Koriyama, H., Mori, M., Gerardo Araujo, R., Maeda, A., Morishita, R., Tamai, K., Kaneda, Y. HIG1, a novel regulator of mitochondrial γ‐secretase, maintains normal mitochondrial function. FASEB J. 26, 2306‐2317 (2012). www.fasebj.org