© 2014 The Authors. Natural killer (NK) cells are an innate lymphoid cell lineage characterized by their capacity to provide rapid effector functions, including cytokine production and cytotoxicity. Here, we identify the Ikaros family member, Aiolos, as a regulator of NK-cell maturation. Aiolos expression is initiated at the point of lineage commitment and maintained throughout NK-cell ontogeny. Analysis of cell surface markers representative of distinct stages of peripheral NK-cell maturation revealed that Aiolos was required for the maturation in the spleen of CD11bhighCD27- NK cells. The differentiation block was intrinsic to the NK-cell lineage and resembled that found in mice lacking either T-bet or Blimp1; however, genetic analysis revealed that Aiolos acted independently of all other known regulators of NK-cell differentiation. NK cells lacking Aiolos were strongly hyper-reactive to a variety of NK-cell-mediated tumor models, yet impaired in controlling viral infection, suggesting a regulatory function for CD27- NK cells in balancing these two arms of the immune response. These data place Aiolos in the emerging gene regulatory network controlling NK-cell maturation and function. Synopsis Aiolos, a member of the Ikaros family of transcription factors, regulates the differentiation of mouse natural killer (NK) cells. NK cells constitutively express Aiolos. Aiolos is required for final stage of NK-cell development in the spleen. Aiolos acts independently of the known regulators of NK-cell maturation. Despite their impaired maturation, NK cells lacking Aiolos show enhanced ability to control tumors. Aiolos, a member of the Ikaros family of transcription factors, regulates the differentiation of mouse natural killer cells.