A20 (also known as TNFAIP3) is a potent anti-inflammatory protein that suppresses many intracellular signaling pathways induced by inflammatory cytokines and bacterial and viral pathogens. The anti-inflammatory function of A20 depends on its modulation of or binding to polyubiquitin chains on key signaling proteins in the nuclear factor-kappa B (NF-kappa B) pathway. To test whether A20 can be used as therapeutic agent in these inflammatory diseases, we prepared a recombinant cell-penetrating form of A20 (TAT-A20) for intracellular delivery and examined its effect on tumor necrosis factor-alpha (TNF alpha)-induced NF-kappa B activation. We observed that TAT-A20 was effectively transduced into cells within 30 min, whereas A20 protein without TAT motive was not. TAT-A20 also inhibited NF-kappa B induction in fibroblasts stimulated with TNF alpha. These results suggest that increasing intracellular level of A20 can be an effective means to suppress NF-kappa B activation and treat inflammatory diseases.