Caspase-mediated cleavage and activation of LIM-kinase 1 and its role in apoptotic membrane blebbing.

ABSTRACT

Actin cytoskeletal reorganization plays a critical role in cell morphological changes, including membrane blebbing during apoptosis. LIM-kinase 1 (LIMK1) regulates actin cytoskeletal reorganization by phosphorylating and inactivating cofilin, an actin filament-depolymerizing and -severing protein. We now report that LIMK1 is cleaved and activated during anti-Fas antibody-induced apoptosis in Jurkat T cells. The cleavage and activation of LIMK1 were blocked by z-DEVD-fmk, an inhibitor for caspase-3 or related proteases, thus indicating that caspase-3-like proteases are responsible for LIMK1 cleavage. The caspase-mediated cleavage of LIMK1 occurs at Asp-240, a site at the N-terminal side of the protein kinase domain, which leads to the production of an N-terminally truncated, constitutively active LIMK1 fragment. Expression of an N-terminally truncated LIMK1 fragment, LIMK1(241-647), induced membrane blebbing in both Jurkat and HeLa cells, with an extent significantly higher than that of wild-type LIMK1. Down-regulation of endogenous LIMK1 expression by small interfering RNA (siRNA) reduced the Fas-induced membrane blebbing in Jurkat cells. These findings suggest that caspase-mediated cleavage and activation of LIMK1 play a role in the membrane bleb formation during apoptosis.