Kinetic analysis of chemotactic peptide-induced actin polymerization in neutrophils.
Definition of the kinetics of ligand-activated actin polymerization in the neutrophil is important for ultimately understanding the mechanisms utilized for regulation of actin polymerization in this non-muscle cell. To better define the kinetics of formyl peptide (fMLP)-induced actin polymerization in neutrophils we determined F-actin content at 5 second intervals after activation of human neutrophils with a range (10(-11)-10(-9) M) of fMLP concentrations. The state of actin polymerization was monitored by quantifying F-actin content with NBD phallacidin binding in both flow cytometric and extraction assays. Results demonstrate three successive kinetic periods of fMLP-induced actin polymerization in neutrophils, a lag period, a 5 second period when rate of polymerization is maximal, and a period of declining rate of actin polymerization as F-actin content approaches a maximum. The duration of the lag period, the maximum rate of polymerization, and the maximum extent of polymerization all depend upon the fMLP concentration. The lag period varies from 0 to 12 seconds and is followed in 5-10 seconds by a 5 second burst of actin polymerization when the rate is as great as 9% increase in F-actin content per second. After the 5 second burst of polymerization, the rate of polymerization rapidly declines. The study defines three distinct kinetic periods of fMLP-induced actin polymerization during which important rate-limiting biochemical events occur. The mechanistic and motile implications of kinetic periods are discussed.
Wang
D H
DH
Department of Cell Biology, University of Alabama, Birmingham School of Medicine.
Berry
K
K
Howard
T H
TH
D000199:Actins / Q000032:analysis / Q000378:metabolism* / Q000493:pharmacokinetics; D002630:Chemotactic Factors / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D006801:Humans; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000032:analysis / Q000187:drug effects / Q000378:metabolism*; D013997:Time Factors
D000199:Actins; D002630:Chemotactic Factors; D009240:N-Formylmethionine Leucyl-Phenylalanine