Malignant cells increase glycolysis and down regulate mitochondrial respiration for ATP production. Mechanisms for respiratory impairment in cancerous cells and their importance for carcinogenesis are not well defined. We found that expression of the respiration-inducing uncoupling protein 3 (UCP3) was normally expressed in murine skin and was greatly decreased in cutaneous malignancies. To better understand the significance of UCP3 in epidermal biology and to test the importance of respiratory changes in cancer development, we generated hemizygous mice expressing a keratin-5 promoter-UCP3 transgene (K5-UCP3). Compared to wild type, K5-UCP3 mice exhibited increased cutaneous mitochondrial respiration, had decreased mitochondrial membrane potential in isolated keratinocytes, and were completely resistant to chemically-induced skin carcinogenesis. We showed that the mechanism of UCP3-dependent cancer protection is most likely not due to increased intracellular heat production or ATP depletion in pre-cancerous cells. Therefore, because hair follicle "bulge" stem cells (bSC) are K5⁺ and progenitors of cutaneous carcinomas, we hypothesized that K5-UCP3 animals were protected from skin carcinogenesis due to alterations in their bSC population. Unlike WT, most (85%) hair follicle bulge regions in K5-UCP3 mice lost biochemical markers of quiescent bSC, but bSC functions were fully intact. Supporting our hypothesis that increased skin turnover protected K5-UCP3 mice from skin cancer; we showed that basal keratinocyte cell cycling was increased 3% in K5-UCP3 skin compared to WT. Moreover, the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) induced similar proliferative responses in both WT and K5-UCP3 skin, but the magnitude of TPA-induced skin thickening was greatly decreased in K5-UCP3 versus WT mice. Together with microarray, histochemical and in vitro morphologic analyses showing that keratinocyte differentiation was sharply increased in K5-UCP3 skin, this implies that UCP3 may increase keratinocyte transit from stem to differentiated daughter cells. Thus, the cancer resistance mechanism in K5-UCP3 mice likely stems from UCP3-induced mitochondrial respiration, which promotes the differentiation and abrogates the tumorigenicity of progenitor keratinocytes. This is the first demonstration in any context that UCP3 blocks carcinogenesis and promotes cellular differentiation. These observations support Warburg's contention that respiratory dysfunction promotes cancer development, and suggest that mitochondrial uncoupling may be a novel target for cancer prevention and treatment. / text