INSULIN AND ATP STIMULATE ACTIN POLYMERIZATION IN U937 CELLS BY A WORTMANNIN-SENSITIVE MECHANISM

ATP and insulin stimulate increases in phosphatidylinositol (3,4,5)-tr isphosphate levels in myeloid-derived U937 cells, Quantification of FI TC-phalloidin binding by fluorescence-activated cell sorting reveals t hat both ATP and insulin stimulate actin polymerization with distincti ve kinetics in U937 cells, The response to ATP is rapid and dose-depen dent with an EC(50) of 200 nM, and is abolished by pre-incubation with the Ca2+ chelator BAPTA-AM. At 800 nM concentration, wortmannin, a po tent inhibitor of phosphoinositide 3-kinase (PI3K), blocks the late, b ut not the early phase of actin polymerization stimulated by 100 nM AT P, Responses elicited by 10 mu g/ml insulin are slower, smaller and mo re transient than responses to ATP, and are inhibited by preincubation with 100 nM wortmannin. Actin polymerization can also be stimulated b y thapsigargin, but not by phorbol ester, providing further evidence f or a role for Ca2+ in actin polymerization. These data implicate disti nct Ca2+ and PI3K-mediated pathways in the regulation of actin polymer ization.