Background <p>How to select muscle-invasive bladder cancer (MIBC) patients who are sensitive to immunotherapy is an unmet medical need. This study aimed to explore the role of immunoproteasome subunits as a novel signature for predicting efficacy of immunotherapy in MIBC.</p> Methods <p>The expression profile of immunoproteasome subunits of MIBC and normal tissues was evaluated from data of The Cancer Genome Atlas (TCGA) and of the Chongqing University Cancer Hospital (CQUCH) cohort. Survival analysis and response to immunotherapy was further explored and compared between immunoproteasome subunits^high and immunoproteasome subunits^low MIBC patients in the TCGA, the CQUCH and the IMvigor210 cohort. The association of the expression of immunoproteasome subunits with immune checkpoint molecules and the tumor immune microenvironment was explored by immunohistochemistry staining and bioinformatic analysis in MIBC of these three cohorts.</p> Results <p>The expression of the immunoproteasome subunits PSMB8, PSMB9 and PSMB10 was significantly upregulated in MIBC. MIBC patients with high expression of immunoproteasome subunits, especially high expression of PSMB9, showed a trend of prolonged overall and progression free survival, which was further significantly improved in response to immunotherapy. Bioinformatics and immunohistochemistry staining revealed a positive correlation of the expression of immunoproteasome subunits with the expression of immune checkpoint molecules, with T cell activation and with T cell-mediated cytotoxicity.</p> Conclusions <p>Immunoproteasome subunits, in particular PSMB9, are immune microenvironment-related molecules of MIBC and are promising signatures for survival prediction in response to immunotherapy of MIBC.</p> Graphical Abstract <p></p>