The retinoic acid-related orphan receptor γ(RORγ) has important roles in development and metabolic homeostasis. Although the biological functions of RORγ have been studied extensively, no ligands for RORγ have been identified, and no structure of RORγ has been reported. In this study,weshowed that hydroxycholesterols promote the recruitment of coactivators by RORγ using biochemical assays. We also report the crystal structures of the RORγ ligand-binding domain bound with hydroxycholesterols. The structures reveal the binding modes of various hydroxycholesterols in the RORγ pocket, with the receptors all adopting the canonical active conformation. Mutations that disrupt the binding of hydroxycholesterols abolish the constitutive activity of RORγ. Our observations suggest an important role for the endogenous hydroxycholesterols in modulating RORγ-dependent biological processes. Copyright © 2010 by The Endocrine Society.