The TGF-b/SOX4 axis and ROS-driven autophagy co-mediate CD39 expression in regulatory T-cells

The ectonucleotidase CD39 on human regulatory T-cells (Treg) is an important immune regulator which is dysregulated in autoimmune diseases and cancer immunosuppression. We here define that CD39 expression on Treg is independent of the Treg-specific transcription factors FOXP3 and HELIOS and promoted by canonical TGF-b- and mTOR-signaling. Furthermore, the TGF-b mediated upregulation of CD39 is counteracted by reactive oxygen species (ROS)-driven autophagy. In line, CD39⁺ peripheral blood Treg constitute a distinct lineage with low autophagic flux and absent ROS production. Patients with rare genetic defects in autophagy show supraphysiological levels of CD39⁺ Treg, validating our observations in vivo. These biological processes rely on a distinct transcriptional program with CD39⁺ Treg expressing low levels of two genes with putative involvement in autophagy, NEFL and PLAC8. Furthermore, the TGF-b downstream transcription factor SOX4 is selectively upregulated in CD39⁺ Treg. Overexpression of SOX4 in Treg strongly increases CD39 expression, while Crispr/Cas9-mediated knockout of SOX4 in Treg has the opposing effect. Thus, we identify a crucial role of SOX4 in immune regulation and provide new insights involving the interplay of tolerogenic cues and autophagy in Treg.