Differential Requirement of Cd8 Enhancers E8I and E8VI in Cytotoxic Lineage T Cells and in Intestinal Intraepithelial Lymphocytes

CD8 expression in T lymphocytes is tightly regulated by the activity of at least six Cd8 enhancers (E8 I -E8 VI ), however their complex developmental stage-, subset-, and lineage-specific interplays are incompletely understood. Here we analyzed ATAC-seq data on the Immunological Genome Project database and identified a similar developmental regulation of chromatin accessibility of a subregion of E8 I , designated E8 I -core, and of E8 VI . Loss of E8 I -core led to a similar reduction in CD8 expression in naïve CD8 + T cells and in IELs as observed in E8 I −/− mice, demonstrating that we identified the core enhancer region of E8 I . While E8 VI −/− mice displayed a mild reduction in CD8 expression levels on CD8SP thymocytes and peripheral CD8 + T cells, CD8 levels were further reduced upon combined deletion of E8 I -core and E8 VI . Moreover, activated E8 I - core −/− E8 VI −/− CD8 + T cells lost CD8 expression to a greater degree than E8 I - core −/− and E8 VI −/− CD8 + T cells, suggesting that the combined activity of both enhancers is required for establishment and maintenance of CD8 expression before and after TCR activation. Finally, we observed a severe reduction of CD4 CTLs among the TCRβ + CD4 + IEL population in E8 I - core −/− but not E8 VI −/− mice. Such a reduction was not observed in Cd8a −/− mice, indicating that E8 I -core controls the generation of CD4 CTLs independently of its role in Cd8a gene regulation. Further, the combined deletion of E8 I -core and E8 VI restored CD4 CTL subsets, suggesting an antagonistic function of E8 VI in the generation of CD4 CTLs. Together, our study demonstrates a complex utilization and interplay of E8 I -core and E8 VI in regulating CD8 expression in cytotoxic lineage T cells and in IELs. Moreover, we revealed a novel E8 I -mediated regulatory mechanism controlling the generation of intestinal CD4 CTLs.