Role of gelsolin interaction with actin in regulation and creation of actin nuclei in chemotactic peptide activated polymorphonuclear neutrophils.

Abstract

In vitro Ca++ activates gelsolin to sever F-actin and form a gelsolin-actin (GA) complex at the + end of F-actin that is not dissociated by ethylene glycol-bis(3-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) but is separated by EGTA + PIP/PIP2. The gelsolin blocks the + end on the actin filament, but the -end of the filament can still initiate actin polymerization. In thrombin activated platelets, evidence suggests that severing of F-actin by gelsolin increases GA complex, creates one -end actin nucleus and one cryptic + end actin nucleus per cut, and then dissociates to yield free + ends to nucleate rapid actin assembly. We examined the role of F-actin severing in creation and regulation of nuclei and polymerization in polymorphonuclear neutrophils (PMNs). At 2-s intervals after formyl peptide (FMLP) activation of endotoxin free (ETF) PMNs, change in GA complex was correlated with change in + end actin nuclei, -end actin nuclei, and F-actin content. GA complex was quantitated by electrophoretograms of proteins absorbed by antigelsolin from cells lysed in 10 mM EGTA, + end actin nuclei as cytochalasin (CD) sensitive and -end actin nuclei as CD insensitive increases in G-pyrenyl actin polymerization rates induced by the same PMNs, and F-actin content by NBDphallacidin binding to fixed cells. Thirty three percent of gelsolin was in GA complex in basal ETF PMNs; from 2-6 s, GA complexes dissociate (low = 15% at 10 s) and sequentially + end nuclei and F-actin content and then -end nuclei increase to a maximum at 10 s. At >40 s GA complex increase toward basal and + end nuclei and F-actin content returned toward basal. These kinetic data show gelsolin regulates availability of + end nuclei and actin polymerization in FMLP. However, absence of an initial increase in GA complex or -end nucleating activity shows FMLP activation does not cause gelsolin to sever F-or to bind G-actin to create cryptic + end nuclei in PMNs; the results suggest the + nucleus formation is gelsolin independent.

Metabolic responses of plant cell culture to hydrodynamic stress

“…F-actin content was measured by fluoromety (Brawley and Robinson, 1985;Li et al, 1994;Deaton et al, 1992;Cao et al, 1992;Howard and Oresajo, 1985). A 0.5-cm3 cell suspension was mixed with 100 cm3 of formaldehyde solution for 30 min.…”

Engagement of the T‐cell antigen receptor by anti‐CD3 monoclonal antibody causes a rapid increase in lymphocyte F‐actin

“…Phosphati-dylinositol4,5 bisphosphate (PIP21 is known to dissociate actin monomers from the sequestering protein, profilin (Goldschmidt-Clermont et al, 1990). PIP2 also dissociates the barbed end blocking protein, gelsolin, from actin (Deaton et al, 1992). The involvement of these mechanisms in the mediation of our observed rise in F-actin was not evaluated.…”

Phospholipase D Regulation by a Physical Interaction with the Actin-Binding Protein Gelsolin

“…Since the rabbit gene encoding gelsolin is not present in GenBank version 86, the four differing amino acids likely represent species differences. Gelsolin catalyzes the severing of actin filaments and is thought to be intimately involved in the control of actin reorganizations through the regulation of filament severing as well as repolymerization (Cunningham, 1992;Deaton et al, 1992;Janmey et al, 1987;McLaughlin et al, 1993;Rozycki et al, 1994;Wegner et al, 1994;Yu et al, 1992).…”