Here we show that miR-21, a microRNA known for its oncogenic activity, is also essential for mediating immune responses against tumor. Knockout of miR-21 in mice slowed the proliferation of both CD4+ and CD8+ cells, reduced their cytokine production and accelerated the grafted tumor growth. Further investigations indicated that miR-21 could activate CD4+ and CD8+ T cells via the PTEN/Akt pathway in response to stimulations. Taken together, these data suggest the key functions of miR-21 in mediating anti-tumor immune response and thereby uncover a bi-directional role of this traditionally known ‘oncomiR’in tumorigenesis. Our study may provide new insights for the design of cancer therapies targeting microRNAs, with an emphasis on the dynamic and possibly unexpected role of these molecules.