T cell anergy is a programmed state of cellular hyporesponsiveness that prevents the propagation of an immune response to self. Thus, anergy is a critical component of self-tolerance. Consequently, the development and maintenance of the anergy program in T cells is tightly controlled by many mechanisms, both cell-intrinsic and cell-extrinsic. Since anergy is important in self-tolerance, it is critical to understand the complex biology that regulates the initiation and regulation of the anergy program. We have found in wild-type (WT) mice that lower expression of the transcription factor Runx1 is associated with anergic T cells, defined by co-expression of the cell-surface markers CD73 and FR4. Furthermore, conditional deletion of Runx1 in T cells using a CD4-Cre produces a higher frequency of T cells that co-express CD73 and FR4. This supports the hypothesis that Runx1 suppresses the development of anergy in CD4+ T cells. CD4-Cre Runx1 cKO mice have a block in T cell maturation and have very few CD4+ T cells in peripheral lymphoid organs. To bypass this maturation defect, and test the role of Runx1 in peripheral CD4+ T cells, we have generated a novel 1:1 mixed bone marrow chimera (BMC) system using Estrogen Receptor (ER)-Cre Runx1 cKO bone marrow mixed with B6.SJL Wild Type (WT) bone marrow. This bypasses the maturation defect by deleting Runx1 in peripheral CD4+ T cells in a tamoxifen inducible manner after they have completed maturation. Furthermore, half the cells are Runx1-sufficient allowing for analysis of T cell-intrinsic effects. In this system, we have found that Runx1 regulates the cell-intrinsic induction of anergy, as increased frequency of co-expression of CD73 and FR4 in peripheral CD4+ T cells is only seen in the ER-Cre Runx1 cKO cells and not the B6.SJL WT cells derived from the same mouse.
Aim 1 of the proposed studies will determine critical gene targets Runx1 regulates to control CD4+ T cell anergy. To do this, candidate genes identified by RNA-sequencing will be analyzed in our novel ER-Cre system, and their role in anergy will functionally assessed. Surprisingly, both the ER-Cre Runx1 cKO and the B6.SJL CD4+ T cells in the tamoxifen treated animals fail to proliferate upon in vitro TCR stimulus, suggesting that Runx1 regulates a cell-extrinsic signaling mechanism controlling tolerance.
Aim 2 will define key mechanisms anergic Runx1-deficient CD4+ T cells use to tolerize wild type CD4+ T cells. To complete this aim we will examine the role of candidate genes in wild type cells. Further investigation of the role of Runx1 in CD4+ T cell anergy will provide insight into how the immune system initiates and maintains self-tolerance.
T cell anergy is a critical component of the immune system that allows for tolerance to self and is a vital mechanism frequently broken in a disease context like autoimmunity. Loss of the transcription factor Runx1 results in an increased frequency of anergic T cells, and these cells are capable of suppressing other T cells in their surrounding environment. This study aims to uncover the mechanisms Runx1 utilizes to regulate immune tolerance and thereby regulate immune responses.
