Thymosin beta 4 sequesters the majority of G-actin in resting human polymorphonuclear leukocytes.
Thymosin beta 4 (T beta 4), a 5-kD peptide which binds G-actin and inhibits its polymerization (Safer, D., M. Elzinga, and V. T. Nachmias. 1991. J. Biol. Chem. 266:4029-4032), appears to be the major G-actin sequestering protein in human PMNs. In support of a previous study by Hannappel, E., and M. Van Kampen (1987. J. Chromatography. 397:279-285), we find that T beta 4 is an abundant peptide in these cells. By reverse phase HPLC of perchloric acid supernatants, human PMNs contain approximately 169 fg/cell +/- 90 fg/cell (SD), corresponding to a cytoplasmic concentration of approximately 149 +/- 80.5 microM. On non-denaturing polyacrylamide gels, a large fraction of G-actin in supernatants prepared from resting PMNs has a mobility similar to the G-actin/T beta 4 complex. Chemoattractant stimulation of PMNs results in a decrease in this G-actin/T beta 4 complex. To determine whether chemoattractant induced actin polymerization results from an inactivation of T beta 4, the G-actin sequestering activity of supernatants prepared from resting and chemoattractant stimulated cells was measured by comparing the rates of pyrenyl-actin polymerization from filament pointed ends. Pyrenyl actin polymerization was inhibited to a greater extent in supernatants from stimulated cells and these results are qualitatively consistent with T beta 4 being released as G-actin polymerizes, with no chemoattractant-induced change in its affinity for G-actin. The kinetics of bovine spleen T beta 4 binding to muscle pyrenyl G-actin are sufficiently rapid to accommodate the rapid changes in actin polymerization and depolymerization observed in vivo in response to chemoattractant addition and removal.
Cassimeris
L
L
Department of Biology, University of Pennsylvania, Philadelphia 19104.
Safer
D
D
Nachmias
V T
VT
Zigmond
S H
SH
D000199:Actins / Q000302:isolation & purification / Q000378:metabolism*; D002630:Chemotactic Factors / Q000494:pharmacology; D002634:Chemotaxis, Leukocyte / Q000502:physiology; D006801:Humans; D046911:Macromolecular Substances; D008954:Models, Biological; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism*; D013347:Subcellular Fractions / Q000737:chemistry; D013947:Thymosin / Q000378:metabolism*
D000199:Actins; D002630:Chemotactic Factors; D046911:Macromolecular Substances; D013947:Thymosin