During endurance training, exercising skeletal muscle experiences severe and repetitive oxygen stress. The primary transcriptional response factor for hypoxic adaptation is hypoxia inducible factor-1 (HIF-1), which upregulates glycolysis and angiogenesis in response to low levels of tissue oxygenation. To examine the role of HIF-1 in endurance training, we have created mice specifically lacking skeletal muscle HIF-1 and subjected them to an endurance training protocol. We found that only wild type mice improve their oxidative capacity, as measured by the respiratory exchange ratio; surprisingly, we found that HIF-1 null mice have already upregulated this parameter without training. Furthermore, untrained HIF-1 null mice have an increased capillary to fiber ratio, and elevated oxidative enzyme activities. These changes correlate with constitutively activated AMP-activated protein kinase in the HIF-1 null muscles. Additionally, HIF-1 null muscles have decreased expression of pyruvate dehydrogenase kinase I, a HIF-1 target that inhibits oxidative metabolism. This data demonstrates that removal of HIF-1 causes an adaptive response in skeletal muscle akin to endurance training, and provides evidence for the suppression of mitochondrial biogenesis by HIF-1 in normal tissue.