NK cells are innate lymphocytes that play an integral role in tumor rejection and viral clearance. Unlike their other lymphocyte counterparts, NK cells have the unique ability to recognize and lyse target cells without prior exposure. However, there are no known NK cell-specific genes that are exclusively expressed by all NK cells. Therefore, identification of NK cell-specific genes would allow a better understanding of why NK cells are unique cytotoxic lymphocytes. From the Immunological Genome (ImmGen) Consortium studies, we identified kruppel-like factor 12 (Klf12), encoding a novel transcription factor, preferentially expressed in C57BL/6 mouse NK cells. KLF12 was dispensable for NK cell development, IFN-γ production, degranulation, and proliferation in Klf12 knockout mice. RNA-sequencing analysis revealed increased expression of Btg3, an antiproliferative gene, in KLF12-deficient NK cells compared with wild-type NK cells. Interestingly, competitive mixed bone marrow chimeric mice exhibited reduced development of KLF12-deficient NK cells, altered IFN-γ production and degranulation, and impairment of NK cell proliferation in vitro and in vivo in response to mouse CMV infection. KLF12-deficient NK cells from bone marrow chimeric mice also expressed higher levels of the IL-21R, which resulted in increased IL-21R signaling and correlated with greater inhibition of NK cell proliferation. Furthermore, IL-21 induced Btg3 expression, which correlated with arrested NK cell maturation and proliferation. In summary, we found that KLF12 regulates mouse NK cell proliferation potentially by regulating expression of Btg3 via IL-21.