The diagram illustrates that the regulatory program of intestinal ROR gamma t positive Treg cells is tightly controlled by two distinct signals during intestinal homeostasis. Signal one, provided by ROR gamma t positive antigen-presenting cells (APCs) via major histocompatibility complex class II-T cell receptor (MHCII-TCR) interactions, and signal two, delivered by other APCs through B7-CD28 costimulation, together shape ROR gamma t positive Treg generation and function. Unexpectedly, signal two antagonizes ROR gamma t positive Treg cells, in contrast to its role in conventional Treg cells. Disruption of signal one leads to loss of ROR gamma t positive Treg cells and intestinal inflammation. Blockade of B7-CD28 costimulation using CTLA4-Ig (Cytotoxic T-Lymphocyte Antigen 4-Immunoglobulin fusion protein, Abatacept) can enhance ROR gamma t positive Treg cell generation and restore intestinal tolerance, but only when signal one from ROR gamma t positive APCs remains intact. These findings highlight the unique, context-dependent regulation of ROR gamma t positive Treg cells and the potential for novel therapeutic strategies that combine preserving signal one with modulating signal two to restore intestinal tolerance.