Abstract The majority of mortality from prostate cancer occurs when the disease progresses to a highly malignant phenotype. Changes in the mitochondrial genome such as mutation, deletion and depletion are common in cancer and can determine advanced phenotype of cancer; however, detailed mechanisms have not been elucidated. We observed that loss of the mitochondrial genome reversibly induced overexpression and activation of proto-oncogenic Ras, especially KRas-4A, responsible for the activation of AKT and ERK leading to advanced phenotype of prostate and breast cancer. Maturation of Ras, possibly prenylation of Ras, was required for Ras activation and was induced by the overexpression of 3- hydroxy-3-methyl-glutaryl-CoA reductase (HMGR), the rate-limiting enzyme of the mevalonate pathway. Hypoxia is known to cause accumulation of intermediate sterols which induces proteasomal degradation of HMGR. Well-differentiated and less aggressive prostate and breast cancer cells with high mitochondrial DNA (mtDNA) content consumed a large amount of oxygen and induced intracellular hypoxia. Loss of mitochondrial genome content reduced oxygen consumption and an increase in oxygen concentration (hypoxic-to-normoxic shift) in the cells. Inhibition of oxygen consumption by the reduction of mitochondrial genome content induced overexpression of HMGR through hypoxic-to-normoxic shift. Subsequently, the endogenous induction of the mevalonate pathway activated Ras that mediates advanced phenotype. Reconstitution of the mitochondrial genome or external hypoxic condition, to reverse the hypoxic-to-normoxic shift, diminished the endogenous induction of the mevalonate pathway and the activation of proto-oncogenic Ras. Reduction of mitochondrial genome content was associated with aggressive phenotype of prostate cancer in vitro cell line model and tissue specimens from prostate cancer patients in vivo. Our results elucidate a coherent mechanism that directly links the changes in mitochondrial genome with the advanced progression of the disease. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1125. doi:1538-7445.AM2012-1125