Active caspase-3 is removed from cells by release of caspase-3-enriched vesicles.

Cleavage of Rho associated Coiled Coil kinase I (ROCK I) by caspase-3 contributes to membrane blebbing. Whether caspase-3 and ROCK I also play a role in the release of membrane vesicles is unknown. Therefore, we transfected a human breast cancer cell line (MCF-7) that is caspase-3 deficient, lacks membrane blebbing, and does not release membrane vesicles, with caspase-3. Cells expressing caspase-3 demonstrate both ROCK I-mediated membrane blebbing, and release of small (400-600nm) membrane vesicles in a ROCK I-independent manner. These membrane vesicles contain caspase-3, and are enriched in caspase-3 activity compared to the releasing cells. Caspase-3-containing vesicles are taken up by untransfected cells but the cells do not show any sign of apoptosis. In conclusion, we show that the release of caspase-3-enriched membrane vesicles and membrane blebbing are two differentially regulated processes. Furthermore, we hypothesize that packaging of caspase-3 into membrane vesicles contributes to cellular homeostasis by the removal of caspase-3, and concurrently, protects the cells' environment from direct exposure to caspase-3 activity.