Mucosa-associated invariant T (MAIT) cells represent a unique population of innate-like T lymphocytes capable of detecting non-peptide antigens in the context of monomorphic antigen-presenting molecules. Due to their abundance in barrier tissues, reactivity to local inflammatory cues, and cytotoxic and regulatory functions, MAIT cells are poised to shape the dynamics of various tumor microenvironments. Growing evidence suggests that MAIT cells can exert protumor and/or antitumor effects in cancers arising from or metastasizing to mucosal tissues. However, MAIT cell roles in bladder cancer (BCa) remains unclear.To begin to identify MAIT cells in BCa, we stained bladder tumor biopsies for T cell receptor (TCR) Vα7.2+ cells. We then refined a human MAIT cell signature, which enabled us to interrogate a bulk RNA sequencing dataset and conduct correlation analyses linking intratumoral MAIT cell abundance and mortality from BCa. To extend our work to an in vivo setting, we employed a clinically relevant mouse model in which Mr1 +/+ B6-MAITCAST (MAIT-sufficient) and Mr1 -/- B6-MAITCAST (MAIT-deficient) mice were exposed to N-butyl-N-(4-hydroxybutyl)nitrosamine, a chemical carcinogen associated with tobacco smoke. In additional experiments, MAIT cells were functionally removed through acetyl-6-formylpterin (Ac-6-FP) administration. Effector and regulatory cell types were phenotyped by flow cytometry, and BCa tumor burden and progression were assessed by MRI and/or H&E and Ki67 staining.TCR Vα7.2+ cells were readily detectable in several BCa biopsies, and our bioinformatic analyses correlated heavier MAIT cell presence in BCa tumors with poorer overall survival. Similarly, we found higher tumor burdens in Mr1+/+ B6-MAITCAST mice than in Mr1-/- or Ac-6-FP-treated animals. Bladder MAIT cells from tumor-bearing mice exhibited phenotypic MAIT17 bias based on transcription factors they harbored along with increased interleukin-17A and tumor necrosis factor-α production capacities upon stimulation. Finally, FoxP3+ regulatory T (Treg) cell frequencies were elevated in Mr1+/+ mouse bladder tumors, likely contributing to an immunosuppressive tumor microenvironment, a finding that could be recapitulated in our transcriptomic studies on human BCa.MAIT cells are abundant in BCa tumor microenvironments where they potentiate Treg cell accumulation and play protumor roles.© Author(s) (or their employer(s)) 2025. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ Group.