Parity-induced changes to mammary epithelial cells control NKT-like cell expansion and mammary oncogenesis

Epigenomic alterations that persist in mammary epithelial cells after pregnancy control cellular responses that induced robust response to pregnancy hormones, and those that impairs c-MYC induced malignant progression. Yet, there is a knowledge gap to whether such pregnancy-induced alterations influences the identify and abundance of specific mammary resident cell populations. Here we utilized single-cell RNA sequencing to define the diversity of epithelial and non-epithelial cells in the post-involuted mammary gland. Our analysis supports the conclusion that a pregnancy cycle induces molecular changes to luminal and basal epithelial populations, which was marked by the up regulation of immune communication signals. Accordingly, we found a specific population of Natural Killer T-like cells that became expanded in healthy, post-involuted mammary glands. Such induction of pregnancy-induced, NKT-like cells were dependent on the expression of the marker CD1d at the surface of epithelial cells, an antigen-presenting molecule that induces NKT maturation. Loss of CD1d expression on mammary epithelial cells, or overall lack of NKT cells, did not altered mammary gland homeostasis, but instead supported the development of mammary oncogenesis in response to cMYC over expression, in a pregnancy-independent manner. Collectively, our findings point to how pregnancy-induced, cell-autonomous changes influence the communication between MECs and normal mammary immune microenvironment, and establishes a causal link between pregnancy-induced epigenetic changes, the immune microenvironment, and mammary oncogenesis.