Negative checkpoint regulators (NCRs) temper the T cell immune response to self-antigens and limit the development of autoimmunity. Unlike all other NCRs that are expressed on activated T lymphocytes, V-type immunoglobulin domain-containing suppressor of T cell activation (VISTA) is expressed on naive T cells, We report an unexpected heterogeneity within the naive I cell compartment in mice, where loss of VISTA disrupted the major quiescent naive T cell subset and enhanced self-reactivity, Agonistic VISTA engagement increased T cell tolerance by promoting antigen-induced peripheral T cell deletion Although a critical player in naive T cell homeostasis, the ability of VISTA to restrain naive T cell responses was lost under inflammatory conditions. VISTA is therefore a distinctive NCR of naive T cells that is critical for steady-state maintenance of quiescence and peripheral tolerance