Abstract Comparative genomics has revealed the rapid expansion of multiple gene families involved in immunity. Members within each gene family often evolved distinct roles in immunity. However, less is known about the evolution of their epigenome and cis -regulation. Here we systematically profile the epigenome of the recently expanded murine Ly49 gene family that mainly encode either inhibitory or activating surface receptors on natural killer cells. We identify a set of cis -regulatory elements (CREs) for activating Ly49 genes. In addition, we show that in mice, inhibitory and activating Ly49 genes are regulated by two separate sets of proximal CREs, likely resulting from lineage-specific losses of CRE activity. Furthermore, we find that some Ly49 genes are cross-regulated by the CREs of other Ly49 genes, suggesting that the Ly49 family has begun to evolve a concerted cis- regulatory mechanism. Collectively, we demonstrate the different modes of cis -regulatory evolution for a rapidly expanding gene family.