Transcriptional regulation of FOXP3 requires integrated activation of both promoter and CNS regions in tumor-induced CD8+ Treg cells

Abstract

T-regulatory cells are an upsurge in the tumor microenvironment and induce immune-evasion. CD4+ Treg cells are well characterized whereas the role of CD8+ Tregs in cancer has recently started to crease attention. Here, we report an augmentation CD8+FOXP3+ Tregs in breast tumor microenvironment. FOXP3, the lineage-specific transcription factor, is a dominant regulator of Treg cell development and function. FOXP3 is induced preferentially by divergent signaling in CD4+ Treg cells. But how FOXP3 is induced and maintained in tumor-CD8+ Tregs is the Cinderella of the investigation. We observed that RUNX3, a CD8+ lineage-specific transcription factor, binds at the FOXP3-promoter to induce its transcription. In addition to promoter activation, involvement of cis-elements CNS1 and CNS2 in the transcriptional regulation of FOXP3 was also evident in these cells. SMAD3 binds to CNS1 region and acts as transcription inducer, whereas GATA3 plays a temporal role in the FOXP3 transcription by differential chromatin modification in CNS regions. In CNS1 region, GATA3 acts as a repressor for FOXP3 in naïve CD8+ T cells. Whereas in CD8+ Tregs, GATA3 binds directly at CNS2 region and persuaded the maintenance of FOXP3. Therefore, the intervention of these concerted transcriptional machinery may have a therapeutic potential in immunotherapy of cancer.

Providence of the CD25+KIR+CD127−FOXP3−CD8+ T‐cell subset determines the dynamics of tumor immune surveillance

“…Interestingly, KIR and CD122 expression was significantly decreased (Figure b) in late‐induced FOXP3 + CD8 + CD25 + CD127 − T cells which expressed a high level of CTLA4 and PD1 as compared to the early‐induced counterpart (Figure c). All these data further discriminate these early‐induced CD8 + CD25 + CD127 − T cells from late‐induced FOXP3 + CD8 + Treg cells . Thus, to the best of our knowledge for the first time, we identified a unique subpopulation of FOXP3 − KIR + CD8 + CD25 + CD127 − T cells in the early tumor microenvironment which had a CD8 + Treg phenotypic signature, which was distinct from CD8 + effector T cells.…”

“…Various evidence suggests that T‐regulatory cells impede immune surveillance and impair immunotherapy . In the tumor microenvironment, FOXP3 + CD8 + Treg cells have the immune suppressive attributes similar to FOXP3 + CD4 + Treg cells . Here, we identified a lineage of FOXP3 − CD25 + CD127 − KIR + CD8 + Treg‐like cells which produce a high level of IFNγ at the early stages of tumor development.…”

“…As we identified a subset of FOXP3 − IFNγ + CD8 + CD25 + CD127 − T cells at the early stages of tumor development which were phenotypically similar to different FOXP3 − CD8 + Treg subsets as well as distinct from effector CD8 + T cells, we next tried to functionally characterize them. Interestingly, it was observed that unlike FOXP3 + CD8 + Tregs, the early‐induced IFNγ + FOXP3 − CD8 + CD25 + CD127 − T cells did not inhibit T‐responder cell proliferation (Figure a) or block Tc‐mediated tumor killing (Figure b). It is to be noted that KIR + CD122 + FOXP3 − CD8 + Treg cells secrete high level of granzyme‐B and kill only HLA‐E‐restricted T FH cells in autoimmune diseases .…”

“…Previous work from our laboratory has shown that FOXP3 + CD8 + Treg cells produce IL10 and TGFβ in the later stage of tumor development . Therefore, we sought to check the cytokine profile of the CD8 + CD25 + CD127 − T cells which were generated in the different stages of the tumor microenvironment.…”

“…Previous work from our laboratory has shown that, like CD4 + FOXP3 + Treg cells, CD8 + FOXP3 + Treg cells are immune suppressive . As we identified a subset of FOXP3 − IFNγ + CD8 + CD25 + CD127 − T cells at the early stages of tumor development which were phenotypically similar to different FOXP3 − CD8 + Treg subsets as well as distinct from effector CD8 + T cells, we next tried to functionally characterize them.…”