Ectodomain shedding of cell surface membrane‐anchoring proteins is an important process in a wide variety of physiological events(1, 2). Tumor necrosis factor α (TNF‐α) converting enzyme (TACE) is the first discovered mammalian sheddase responsible for cleavage of several important surface proteins, including TNF‐α, TNF p75 receptor, L‐selectin, and transforming growth factor‐α. Phorbol myristate acetate (PMA) has long been known as a potent agent to enhance ectodomain shedding. However, it is not fully understood how PMA activates TACE and induces ectodomain shedding. Here, we demonstrate that PMA induces both reactive oxygen species (ROS) generation and TNF p75 receptor shedding in Mono Mac 6 cells, a human monocytic cell line, and l‐selectin shedding in Jurkat T‐cells. ROS scavengers significantly attenuated PMA‐induced TNF p75 receptor shedding. Exogenous H2O2 mimicked PMA‐induced enhancement of ectodomain shedding, and H2O2‐induced shedding was blocked by TAPI, a TACE inhibitor. Furthermore, both PMA and H2O2 failed to cause ectodomain shedding in a cell line that lacks TACE activity. By use of an in vitro TACE cleavage assay, H₂O₂ activated TACE that had been rendered inactive by the addition of the TACE inhibitory pro‐domain sequence. We presume that the mechanism of TACE activation by H2O2 is due to an oxidative attack of the pro‐domain thiol group and disruption of its inhibitory coordination with the Zn⁺⁺ in the catalytic domain of TACE. These results demonstrate that ROS production is involved in PMA‐induced ectodomain shedding and implicate a role for ROS in other shedding processes.