Sirtuin-3 (Sirt3) is an essential metabolic regulatory enzyme that plays an important role in mitochondrial metabolism, but its role in bone marrow and skeletal homeostasis remains largely unknown. In this study, we hypothesize that increased expression of Sirt3 plays a role in skeletal aging. Using mice that overexpress Sirt3 [i.e., Sirt3 transgenic (Sirt3Tg)], we show that Sirt3 is a positive regulator of adipogenesis and osteoclastogenesis and a negative regulator of skeletal homeostasis. Sirt3Tg mice exhibited more adipocytes in the tibia compared with control mice. Bone marrow stromal cells (BMSCs) from Sirt3Tg mice displayed an enhanced ability to differentiate into adipocytes compared with control BMSCs. We found a 2.5-fold increase in the number of osteoclasts on the bone surface in Sirt3Tg mice compared with control mice (P < 0.03), and increased osteoclastogenesis in vitro. Importantly, Sirt3 activates the mechanistic target of rapamycin (mTOR) pathway to regulate osteoclastogenesis. Sirt3Tg male mice exhibited a significant reduction in cortical thickness at the tibiofibular junction (P < 0.05). In summary, Sirt3 activity in bone marrow cells is associated with increased adipogenesis, increased osteoclastogenesis through activation of mTOR signaling, and reduced bone mass. Interestingly, Sirt3 expression in bone marrow cells increases during aging, suggesting that Sirt3 promotes age-related adipogenesis and osteoclastogenesis associated with bone loss. These findings identify Sirt3 as an important regulator of adipogenesis and skeletal homeostasis in vivo and identify Sirt3 as a potential target for the treatment of osteoporosis.