UCSF

T cells undergo dynamic and rapid changes in gene expression and protein output in response to cognate antigen, processes aided by post-transcriptional regulation. RNA binding proteins (RPBs) are important trans factors that bind to the 3’ untranslated region (3’UTR) of transcripts and modulate mRNA degradation and translation. Loss of function of these proteins, as well as mutations in their cis-regulatory binding sites, can alter T function and development and lead to immune-mediated diseases. While studies have characterized the network of RBPs present in T cells, as well as binding profiles of individual RBPs, the cis-regulatory landscape of the T cell transcriptome is yet to be defined. This body of work addresses this gap by developing a biochemical assay to map the RBP binding profile in T cells and identify post-transcriptional cis-regulatory regions that regulate protein expression and T cell function. Using our technique, GCLiPP, we identified biochemically shared binding sites across conserved regions of human and mouse 3’UTRs that govern transcript stability. Additional dissections of predicted RBP binding sites in several 3’UTRs of immune-related genes revealed cis-regulatory regions critical for modulating protein expression. We performed further analysis on CD69, a negative regulator of T cell egress, and identified a conserved destabilizing region within the 3’UTR. Absence of this region, as well as the whole 3’UTR, increased CD69 transcript half-life and led to higher CD69 expression as well as more CD69+ T cells in mouse and human. The post-transcriptional circuitry within Cd69 3’UTR had a profound effect on T cell migration, as mice with homozygous deletion of this region showed impaired thymic egress. Taken together, our studies provide a resource for mapping post-transcriptional cis-elements in the T cell transcriptome and demonstrate its utility for discovering functional cis-regulatory regions that contain variants associated with human immune-mediated diseases.