Mapping REV-ERBα’s Protein-Protein Interactions to Understand TH17 Cell Regulation and Inflammation 2258028

Abstract

TH17 cells are a subset of CD4+ T cells that mediate autoimmune and chronic inflammatory pathology. Our lab has previously demonstrated that REV-ERBα, a member of the nuclear receptor superfamily of ligand-regulated transcription factors, represses TH17 development in vitro and protects against relevant models of disease in vivo. While REV-ERBα’s target genes and binding sites have been identified across multiple tissues, little is known about the transcriptional machinery that drives its function. We coupled MiniTurboID proximity labeling with label-free proteomics to 1) identify key proteins that facilitate REV-ERBα-mediated gene repression in primary, murine TH17 cells and 2) further understand how ligands modulate this process. Furthermore, we have conducted a pooled in vivo RNAi screen to examine our proteomics hits in a physiologically relevant context. Specifically, we utilized Il17a reporter REV-ERB deficient and sufficient T-cells along with shRNAs targeting our identified REV-ERBα interactors in a mouse model of colitis. Comparison of the REV-ERBα interactome induced by the presence and absence of endogenous ligand binding reveals potential canonical and non-canonical mechanisms of repression. Additionally, comparison of the shRNAs enriched in colon cells expressing high versus low levels of IL-17A have elucidated which REV-ERBα interactors regulate TH17 pathogenicity in a REV-ERB-dependent manner. Ultimately, these efforts aim to bolster our understanding of the REV-ERBs, nuclear receptor biology, and TH17 pathogenesis–all of which may inform the development of focused therapeutics for treatment of autoimmune and chronic inflammatory diseases. NA Basic Autoimmunity (BA)