The molecular circuits controlling human T follicular regulatory cell development

PROJECT ABSTRACT GC responses are steered toward pathogen-recognition and away from self-reactivity by T follicular regulatory (Tfr) cells. The molecular means by which Tfr cells exercise control are unknown. GCs are deranged and Tfr subsets are altered in lymph nodes excised from common variable immunodeficiency (CVID) patients. CVID is a genetic disease caused by mutations in immune critical genes like CTLA4, NFKB1, PIK3CD or PIK3R1 and diagnosed based upon poor vaccine responses and a proclivity for autoantibody production. The relevance of Tfr cell defects to CVID pathogenesis is unclear. The long-range goal of the proposed work is to determine how GCs are controlled in health and how governance loss promotes immune deficiencies and autoimmune diseases. The objective of this grant application is to establish which lineage lymph node Tfr cells belong to in humans and how lineage-specific Tfr cell developmental aberrations contribute to autoimmune pathologies. The working hypothesis is that the Tfr cell pool encompasses both “natural” nTfr cells that descended from Tregs and iTfr cells that are induced from T follicular helper (Tfh) cells. Whereas nTfrs express autoreactive T-cell receptors and enforce self-tolerance, iTfrs arrive weeks into a GC response to amplify affinity maturation and ultimately quinch the reaction. Based upon altered circulating iTfr to nTfr ratios in CVID patients, we anticipate counterpart cells in their paired lymph nodes will display altered frequencies, locations and repertoires. Our rationale is that by understanding which GC response elements are controlled by iTfrs and nTfrs, we can develop new diagnostic and therapeutic opportunities for patients with GC-dysfunction disease including, but not limited to, CVID. Our specific aims will test the following hypotheses: Aim 1: iTfr and nTfr cells can be clonally distinguished and functionally described in lymph nodes and blood from controls and CVID patients Aim 2: iTfr and nTfr cells can be localized within healthy and CVID lymph node sections and Aim 3: CVID Tfr defects and CVID-associated GC dysregulation can be reproduced in genetically edits tonsil organoids. The contribution is significant because the molecular underpinnings of human GC regulation are unknown are of great consequence to patients with autoimmune and immunodeficiency diseases. The proposed work is innovative because we propose defining protein and gene expression of single human cells in their native, tissue-resident spatial context with a suite of modern, high-dimensional technologies uniquely suited to our topic. Further, we are verifying our findings in rare excisional lymph node biopsies of CVID patients using high-dimensional imaging and disease modeling in genetically edited tonsil organoids.