Effects of CapG overexpression on agonist-induced motility and second messenger generation.

Actin modulating proteins that bind poly- phosphoinositides, such as phosphatidylinositol 4, 5-bisphosphate (PIPE), Can potentially participate in receptor signaling by restructuring the membrane cytoskeleton and modulating second messenger gener- ation through the phosphoinositide cycle. We examined these possibilities by overexpressing CapG, an actin filament end capping, Ca 2÷- and polyphosphoinostide- binding protein of the gelsolin family. High level tran- sient overexpression decreased actin 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 con- tent or organization. Nevertheless, it promoted a dose- dependent increase in rates of wound healing and chemotaxis. The motile phenotype was similar to that oobserved with gelsolin overexpression, which in addi- tion to capping, also severs and nucleates actin illa- ments. CapG overexpressing clones are more respon- sive to platelet-derived growth factor than control- transfected clones. They form more circular dorsal membrane ruffles, have higher phosphoinositide turn- over, inositol 1,4,5-trisphosphate generation and Ca 2÷ signaling. These responses are consistent with en- hanced PLC'y activity. Direct measurements of PIP2 mass showed that the CapG effect on PLC'y was not due primarily to an increase in the PIP2 substrate con- centration. The observed changes in cell motility and membrane signaling are consistent with the hypothesis that PIP2-binding actin regulatory proteins modulate phosphoinositide turnover and second messenger gen- eration in vivo. We infer that CapG and related pro- teins are poised to coordinate membrane signaling with actin filament dynamics following cell stimulation.