Over the past several years, SIRT5 has attracted considerable attention in metabolic regulation. However, the function of SIRT5 in immune cells is poorly understood. We discovered that proteome and network analysis revealed SIRT5 was important in T cell receptor signaling pathways. Additionally, we determined that a deficiency in SIRT5 induced T cell activation. Furthermore, our results demonstrated that SIRT5 played a pivotal role in regulating the differentiation of regulatory T (Treg) cells and T-helper 1 (Th1) cells. An imbalance in the lineages of immunosuppressive Treg cells and the inflammatory Th1 subsets of helper T cells leads to the development of inflammatory diseases and cancer. Furthermore, we found that Sirt5 knockout mice were resistant to AOM and DSS-induced colitis-associated colorectal tumorigenesis.