The actin cytoskeleton facilitates complement-mediated activation of cytosolic phospholipase [A.sub.2]

Cytosolic PL[A.sub.2-[alpha]] (cPL[A.sub.2]) and metabolites of arachidonic acid (AA) are key mediators of complement-dependent glomerular epithelial cell (GEC) injury. Assembly of C5b-9 increases cytosolic [Ca.sup.2+] concentration and results in transactivation of receptor tyrosine kinases and activation of PLC-[gamma]1 and the 1,2-diacylglycerol (DAG)-PKC pathway. [Ca.sup.2+] and PKC are essential for membrane association and increased catalytic activity of cPL[A.sub.2]. This study addresses the role of the actin cytoskeleton in cPL[A.sub.2] activation. Depolymerization of F-actin by cytochalasin D or latrunculin B reduced complement-dependent [[sup.3]H]AA release, as well as the complement-induced increase in cPL[A.sub.2] activity. These effects were due to inhibition of [[sup.3]H]DAG production and PKC activation, implying interference with PLC. Complement-dependent [[sup.3]H]AA release was also reduced by jasplakinolide, a compound that stabilizes F-actin and organizes actin filaments at the cell periphery, and calyculin A, which induces condensation of actin filaments at the plasma membrane. The latter drugs did not affect [[sup.3]H]DAG production, suggesting their inhibitory actions were downstream of PKC. Neither cytnchalasin D, latrunculin B, nor calyculin A affected association of cPL[A.sub.2] with microsomal membranes, and cytochalasin D and latrunculin B did not alter the localization of the endoplasmic reticulum. Stable transfection of constitutively active RhoA induced formation of stress fibers, stabilized F-actin, and attenuated the complement-induced increase in [[sup.3]H]AA. Thus in GEC, cPL[A.sub.2] activation is dependent, in part, on actin remodeling. By regulating complement-mediated activation of cPL[A.sub.2], the actin cytoskeleton may contribute to the pathophysiology of GEC injury. inflammation: lipid mediators; protein kinases; signal transduction