AUTHOR=Gülich Alexandra Franziska , Preglej Teresa , Hamminger Patricia , Alteneder Marlis , Tizian Caroline , Orola Maria Jonah , Muroi Sawako , Taniuchi Ichiro , Ellmeier Wilfried , Sakaguchi Shinya 

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

JOURNAL=Frontiers in Immunology

VOLUME=10

YEAR=2019

URL=https://www.frontiersin.org/journals/immunology/articles/10.3389/fimmu.2019.00409

DOI=10.3389/fimmu.2019.00409

ISSN=1664-3224

ABSTRACT=<p>CD8 expression in T lymphocytes is tightly regulated by the activity of at least six <italic>Cd8</italic> enhancers (E8<sub>I</sub>-E8<sub>VI</sub>), 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<sub>I</sub>, designated E8<sub>I</sub>-core, and of E8<sub>VI</sub>. Loss of E8<sub>I</sub>-core led to a similar reduction in CD8 expression in naïve CD8<sup>+</sup> T cells and in IELs as observed in <italic>E8</italic><sub><italic>I</italic></sub><sup>−/−</sup> mice, demonstrating that we identified the core enhancer region of E8<sub>I</sub>. While <italic>E8</italic><sub><italic>VI</italic></sub><sup>−/−</sup> mice displayed a mild reduction in CD8 expression levels on CD8SP thymocytes and peripheral CD8<sup>+</sup> T cells, CD8 levels were further reduced upon combined deletion of E8<sub>I</sub>-core and E8<sub>VI</sub>. Moreover, activated <italic>E8</italic><sub><italic>I</italic></sub><italic>-</italic>core<sup>−/−</sup><italic>E8</italic><sub><italic>VI</italic></sub><sup>−/−</sup> CD8<sup>+</sup> T cells lost CD8 expression to a greater degree than <italic>E8</italic><sub><italic>I</italic></sub><italic>-</italic>core<sup>−/−</sup> and <italic>E8</italic><sub><italic>VI</italic></sub><sup>−/−</sup> CD8<sup>+</sup> 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<sup>+</sup>CD4<sup>+</sup> IEL population in <italic>E8</italic><sub><italic>I</italic></sub><italic>-</italic>core<sup>−/−</sup> but not <italic>E8</italic><sub><italic>VI</italic></sub><sup>−/−</sup> mice. Such a reduction was not observed in <italic>Cd8a</italic><sup>−/−</sup> mice, indicating that E8<sub>I</sub>-core controls the generation of CD4 CTLs independently of its role in <italic>Cd8a</italic> gene regulation. Further, the combined deletion of E8<sub>I</sub>-core and E8<sub>VI</sub> restored CD4 CTL subsets, suggesting an antagonistic function of E8<sub>VI</sub> in the generation of CD4 CTLs. Together, our study demonstrates a complex utilization and interplay of E8<sub>I</sub>-core and E8<sub>VI</sub> in regulating CD8 expression in cytotoxic lineage T cells and in IELs. Moreover, we revealed a novel E8<sub>I</sub>-mediated regulatory mechanism controlling the generation of intestinal CD4 CTLs.</p>