MODULATION OF GELSOLIN FUNCTION - ACTIVATION AT LOW PH OVERRIDES CA2+REQUIREMENT

The activation of gelsolin by calcium has been postulated to be involv ed in the receptor-mediated reorganization of the actin cytoskeleton, but cytoskeletal reorganization can also occur in cells with intracell ular Ca2+ clamped at nanomolar levels. Fluorescence measurements using Fura-2 show that at pH 7.4, the Ca2+ requirement for gelsolin activat ion in vitro is higher than previously reported, with half-maximal act ivation of severing and nucleation occurring at 10 muM Ca2+. The Ca2requirement for gelsolin activity decreases at more acid pH and is app roximately 3 muM at pH 6.5. At pH below 6.0, gelsolin no longer requir es Ca2+ for activity and severs actin filaments, binds two actin monom ers, and nucleates filament formation in EGTA-containing solutions. Th e pH-activated severing activity is inhibited by mixed lipid vesicles containing phosphatidylinositol 4,5-bisphosphate. A Ca2+-sensitive fra gment consisting of the first 135 amino acids of human cytoplasmic gel solin also demonstrates severing activity at pH <6.0 in the absence of Ca2+. In contrast, the gelsolin homologs severin and villin maintain Ca2+ regulation of severing activity at low pH. These differences sugg est that activation of gelsolin at low pH cannot be explained merely b y destabilization of F-actin. The difference in diffusion constants of gelsolin measured at pH 5.5 and 6.5, as determined by dynamic light s cattering, suggests that the molecule undergoes a shape change similar to that reported upon binding Ca2+ at neutral pH. These results sugge st a mechanism by which gelsolin may be activated in vivo under condit ions where Ca2+ transients do not occur.