Self-Peptides Bound to MHC Class II in T Cell Selection

Abstract Regulatory T (Treg) cells, serving as life-long guardians of the immune system, are distinguished by constitutive expression of transcription factor (TF) Foxp3, playing a critical role in their differentiation, function and fitness. Our previous studies showed that Foxp3 expression is stable in fully differentiated Treg cells, while recently generated Treg cells can lose Foxp3 expression. Furthermore, Treg cell functionality and Foxp3 expression can be compromised in severe disease settings. Although the unresolved issue of resilience vs. vulnerability of Foxp3-dependent Treg functional program is of major basic and clinical significance, presently the role for Foxp3 protein in functional and transcriptional program of differentiated Treg cells in vivo remains unknown. Our preliminary studies of induced degradation of Foxp3 protein in vivo using novel optimized Foxp3 auxin-degron model raise the possibility of a remarkable resilience of ‘mature’ Foxp3-dependent Treg cell function and gene regulatory network (GRN) in adulthood, but not of ‘immature’ ones in early life. Based on our published and preliminary studies we hypothesize that during Treg differentiation, initially vulnerable Foxp3 dependent GRN approaches a temporally self-sustained (attractor) state over a surprisingly long timescale due to a requirement for persistent Foxp3 activity for this ‘maturation’. Upon assumption of this state, its resilience ensures extended stability of Treg function even upon the loss of Foxp3 expression. In this proposal we will test this hypothesis by elucidating Foxp3 GRN underlying Treg cell function, dynamics of its establishment and its resilience to Foxp3 loss in physiologic and inflammatory settings using genetic gain- and loss-of-function approaches. The following Specific Aims will be pursued: 1) elucidate a role of Foxp3 in the functional program of Treg cells and its resilience in early life and adulthood under physiologic conditions and upon inflammatory challenges; 2) investigate dynamic of Foxp3-dependent regulation of gene expression and chromatin in Treg cells using chemogenetic gain- and loss-of-function approaches; 3) elucidate mechanisms of resilience of Foxp3-dependent gene expression and functional programs using CRISPR and CRISPRi/a Perturb-seq approaches. These studies will provide mechanistic, gene-regulation level framework for understanding of Treg cell differentiation and function in health and disease and inform novel approaches for therapeutic targeting of Treg cells and development of adoptive Treg cell therapies for autoimmune and inflammatory disorders.