Regulation of membrane phospholipid asymmetry by Notch-mediated flippase expression controls the number of intraepithelial TCRαβ⁺CD8αα⁺ T cells

by Chieko Ishifune, Shin-ichi Tsukumo, Yoichi Maekawa, Katsuto Hozumi, Doo Hyun Chung, Chihiro Motozono, Sho Yamasaki, Hiroyasu Nakano, Koji Yasutomo

Intestinal intraepithelial lymphocytes (IELs) expressing CD8αα on αβ T cells (TCRαβ⁺CD8αα⁺ IELs) have suppressive capabilities in enterocolitis, but the mechanism that maintains homeostasis and cell number is not fully understood. Here, we demonstrated that the number of TCRαβ⁺CD8αα⁺ IELs was severely reduced in mice lacking recombination signal binding protein for immunoglobulin kappa J region (Rbpj) or Notch1 and Notch2 in T cells. Rbpj-deficient TCRαβ⁺CD8αα⁺ IELs expressed low levels of Atp8a2, which encodes a protein with flippase activity that regulates phospholipid asymmetry of plasma membrane such as flipping phosphatidylserine in the inner leaflet of plasma membrane. Rbpj-deficient TCRαβ⁺CD8αα⁺ IELs cannot maintain phosphatidylserine in the inner leaflet of the plasma membrane. Furthermore, depletion of intestinal macrophages restored TCRαβ⁺CD8αα⁺ IELs in Rbpj-deficient mice, suggesting that exposure of phosphatidylserine on the plasma membrane in Rbpj-deficient TCRαβ⁺CD8αα⁺ IELs acts as an “eat-me” signal. Together, these results revealed that Notch–Atp8a2 is a fundamental regulator for IELs and highlighted that membrane phospholipid asymmetry controlled by Notch-mediated flippase expression is a critical determinant in setting or balancing the number of TCRαβ⁺CD8αα⁺ IELs.