Wiskott-Aldrich syndrome protein (WASP) links T-cell receptor (TCR) signaling to the actin cytoskeleton. WASP is normally protected from degradation by the Ca++-dependent protease calpain and by the proteasome because of its interaction with the WASP-interacting protein.Objective:
We investigated whether WASP is degraded after TCR ligation and whether its degradation downregulates F-actin assembly caused by TCR ligation.Methods:
Primary T cells, Jurkat T cells, and transfected 293T cells were used in immunoprecipitation experiments. Intracellular F-actin content was measured in splenic T cells from wild-type, WASP-deficient, and c–Casitas B-lineage lymphoma (Cbl)-b–deficient mice by using flow cytometry. Calpeptin and MG-132 were used to inhibit calpain and the proteasome, respectively.Results:
A fraction of WASP in T cells was degraded by calpain and by the ubiquitin-proteasome pathway after TCR ligation. The Cbl-b and c-Cbl E3 ubiquitin ligases associated with WASP after TCR signaling and caused its ubiquitination. Inhibition of calpain and lack of Cbl-b resulted in a significantly more sustained increase in F-actin content after TCR ligation in wild-type T cells but not in WASP-deficient T cells.Conclusion:
TCR ligation causes WASP to be degraded by calpain and to be ubiquitinated by Cbl family E3 ligases, which targets it for destruction by the proteasome. WASP degradation might provide a mechanism for regulating WASP-dependent TCR-driven assembly of F-actin.