p66Shc regulates vesicle-mediated secretion in mast cells by affecting F-actin dynamics.

The extracellular vesicular compartment has emerged as a novel system of intercellular communication; however, the mechanisms involved in membrane vesicle biogenesis and secretion are as yet unclear. Among immune cells releasing membrane vesicles-mast cells that reside near tissues exposed to the environment-are master modulators of immune responses. Here, we have addressed the role of p66Shc, a novel regulator of mast cell activation and homeostasis, in the dynamic reorganization of the actin cytoskeleton that is associated with morphological changes during secretion. We show that p66Shc is recruited as a complex with the lipid phosphatase SHIP1 to the F-actin skeleton and impairs antigen-dependent cortical F-actin disassembly and membrane ruffling through the inhibition of Vav and paxillin phosphorylation. We also show that in addition to acting as a negative regulator of antigen-dependent mast cell degranulation, p66Shc limits the basal release of granule contents by inhibiting microvesicle budding from the plasma membrane and piecemeal degranulation. These findings identify p66Shc as a critical regulator of actin dynamics in mast cells, providing a basis for understanding the molecular mechanisms involved in vesicle-mediated secretion in these cells.