AbstractMucosal-associated invariant T (MAIT) cells express an invariant TRAV1/TRAJ33 TCR-α chain and are restricted to the MHC-I-like molecule, MR1. Whether MAIT cell development depends on this invariant TCR-α chain is unclear. Here we generate Traj33-deficient mice and show that they are highly depleted of MAIT cells; however, a residual population remains and can respond to exogenous antigen in vitro or pulmonary Legionella challenge in vivo. These residual cells include some that express Trav1⁺ TCRs with conservative Traj-gene substitutions, and others that express Trav1^- TCRs with a broad range of Traj genes. We further report that human TRAV1-2^- MR1-restricted T cells contain both MAIT-like and non-MAIT-like cells, as judged by their TCR repertoire, antigen reactivity and phenotypic features. These include a MAIT-like population that expresses a public, canonical TRAV36⁺ TRBV28⁺ TCR. Our findings highlight the TCR diversity and the resulting potential impact on antigen recognition by MR1-restricted T cells.