Sirtuins are highly conserved NAD+-dependent enzymes that are capable of removing a wide range of lipid lysine acyl-groups from protein substrates in a NAD+-dependent manner. These NAD+-dependent activities enable sirtuins to monitor cellular energy status and modulate gene transcription, genome stability, and energy metabolism in response to environmental signals. Consequently, sirtuins are important for cell survival, stress resistance, proliferation, and differentiation. In recent years, sirtuins are increasingly recognized as crucial regulators of stem cell biology in addition to their well-known roles in metabolism and aging. This review article highlights our current knowledge on sirtuins in stem cells, including their functions in pluripotent stem cells, embryogenesis, and development as well as their roles in adult stem cell maintenance, regeneration, and aging.