Endothelial cells (ECs) require glycolysis for proliferation and migration during angiogenesis; however, the necessity for the mitochondrial respiratory chain during angiogenesis is not known. Here we report that inhibition of respiratory chain complex III impairs proliferation, but not migration of ECs in vitro by decreasing the NAD+/NADH ratio. To determine whether mitochondrial respiration is necessary for angiogenesis in vivo, we conditionally ablate a subunit of the respiratory chain complex III (QPC) in ECs. Loss of QPC decreases respiration, resulting in diminished EC proliferation, and impairment in retinal and tumor angiogenesis. Loss of QPC does not decrease genes associated with anabolism or nucleotides levels in ECs, but diminishes amino acid levels. Our findings indicate that mitochondrial respiration is necessary for angiogenesis, and that the primary role of mitochondria in ECs is to serve as biosynthetic organelles for cell proliferation.