Towards molecular mechanisms for pathogenic Th17 cell differentiation in ocular autoimmunity

Abstract

Abstract CD4+ T helper (Th) cells play critical roles in coordinating proper immune responses by differentiating into effector subtypes specialized against different types of pathogens, while dysregulated Th cells can drive autoimmune disease by triggering inappropriate immune responses. Pathogenic Th17 (pTh17) cells, which combine behaviors of classic Th1 and Th17 effector cells by producing both interferon-gamma (IFNg) and interleukin-17A (IL17A), have been implicated in autoimmune diseases. Our goal is to map regulatory networks and identify novel regulators underlying pTh17 differentiation and activation using the murine model experimental autoimmune uveitis (EAU), in which ocular autoimmunity is driven by pTh17 cells. To track pTh17 differentiation, we bred a double reporter mouse with IL17A-cre/tdTomato-flox tracer and IFNg-YFP. We sorted both Tomato+YFP+ and Tomato+YFP- CD4+ T cells from eyes and spleens after EAU induction for single-cell RNA-seq. Clustering of single-cell transcriptomes revealed distinct transcriptomes between eyes and spleen. In the eye, we identified clusters similar to pTh17 and non-pathogenic (np)Th17 populations defined in a myelin autoimmunity model. The pTh17 cluster contained both double positive and Tomato single positive cells, suggesting a heterogeneous pathogenic population. In contrast, the npTh17 cluster only contained single positives. Differential gene expression analysis showed that pathogenic and non-pathogenic clusters utilize distinct sets of transcription factors. We are further characterizing epigenetic differences between pTh17 and npTh17 to better understand how distinct regulatory networks shape pTh17 differentiation. Supported by the NEI Intramural Research Program fund (ZIAEY000569-01)