EFFECTS OF CAPG OVEREXPRESSION ON AGONIST-INDUCED MOTILITY AND 2ND-MESSENGER GENERATION

Actin modulating proteins that bind polyphosphoinositides, such as pho sphatidylinositol 4, 5-bisphosphate (PIP2), can potentially participat e in receptor signaling by restructuring the membrane cytoskeleton and modulating second messenger generation through the phosphoinositide c ycle. We examined these possibilities by overexpressing CapG, an actin filament end capping, Ca2+- and polyphosphoinostide-binding protein o f the gelsolin family. High level transient overexpression decreased a ctin filament staining in the center of the cells but not in the cell periphery. Moderate overexpression in clonally selected cell lines did not have a detectible effect on actin filament content or organizatio n. Nevertheless, it promoted a dose-dependent increase in rates of wou nd healing and chemotaxis. The motile phenotype was similar to that oo bserved with gelsolin overexpression, which in addition to capping, al so severs and nucleates actin filaments. CapG overexpressing clones ar e more responsive to platelet-derived growth factor than control-trans fected clones. They form more circular dorsal membrane ruffles, have h igher phosphoinositide turnover, inositol 1,4,5-trisphosphate generati on and Ca2+ signaling. These responses are consistent with enhanced PL C gamma activity. Direct measurements of PIP2 mass showed that the Cap G effect on PLC gamma was not due primarily to an increase in the PIP2 substrate concentration. The observed changes in cell motility and me mbrane signaling are consistent with the hypothesis that PIP2-binding actin regulatory proteins modulate phosphoinositide turnover and secon d messenger generation in vivo. We infer that CapG and related protein s are poised to coordinate membrane signaling with actin filament dyna mics following cell stimulation.