BETA2/NeuroD, a basic helix-loop-helix transcription factor, is a key regulator of pancreatic islet morphogenesis and insulin gene transcription. Here we report for the first time that the BETA2/NeuroD protein can permeate several cells, including pancreatic islets, due to an arginine- and lysine-rich protein transduction domain sequence in its structure. The BETA2/NeuroD protein was transduced in a dose-dependent manner up to 1 micromol/l. Transduced BETA2/NeuroD functions similarly to endogenous BETA2/NeuroD: it binds to the insulin promoter and activates its expression. We also investigated the mechanism of BETA2/NeuroD protein transduction. The BETA2/NeuroD protein penetrated cells by macropinocytosis and was released from endosomes homogeneously in cytoplasm and nuclei. These data suggest that BETA2/NeuroD protein transduction could be a safe and valuable strategy for enhancing insulin gene transcription without requiring gene transfer technology.