Ca2+-dependent myosin II activation is required for uropod retraction during neutrophil migration

J Cell Sci. 2000 Apr:113 ( Pt 7):1287-98. doi: 10.1242/jcs.113.7.1287.

Abstract

Buffering of intracellular Ca2+ transients in human neutrophils leads to reduced motility due to defective uropod detachment on fibronectin and vitronectin-coated surfaces. Since one potential target of a rise in [Ca2+]i is the activation of myosin II, we characterized the role of myosin II during motility. Treatment of neutrophils with a myosin inhibitor (2,3-butanedione monoxime), or myosin light chain kinase inhibitors (ML-7, ML-9, or KT5926) resulted in impaired uropod retraction and a dose-dependent decrease in chemokinesis following stimulation with N-formyl-Met-Leu-Phe (fMLP). Treatment with ML-9 resulted in a redistribution of F-actin and talin to the non-retracted uropods, mimicking the redistribution observed during [Ca2+]i buffering. Impairment of uropod retraction and redistribution of F-actin and talin by myosin II inhibition was only observed on adhesive substrates such as fibronectin and not on poorly adhesive substrates such as human serum-coated glass. At higher concentrations of ML-9, cell polarization was inhibited and pseudopod extension occurred radially. Using an antibody specific for serine 19-phosphorylated regulatory light chain of myosin II, regions of activated myosin II were found at the leading edge as well as the uropod in motile fMLP-stimulated cells. [Ca2+]i depletion caused a 50% decrease in the level of serine 19-phosphorylated myosin II suggesting that activation of myosin II by intracellular Ca2+ transients may be an essential step in establishing a polarized pseudopod and providing the force required for uropod retraction during PMN motility on adhesive surfaces.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Actins / metabolism
  • Azepines / pharmacology
  • Buffers
  • Calcium / physiology*
  • Cell Adhesion / drug effects
  • Cell Adhesion / physiology
  • Cell Movement / drug effects
  • Cell Movement / physiology*
  • Cell Polarity / drug effects
  • Chemotaxis, Leukocyte / drug effects
  • Chemotaxis, Leukocyte / physiology
  • Dose-Response Relationship, Drug
  • Enzyme Inhibitors / pharmacology
  • Humans
  • Myosin-Light-Chain Kinase / antagonists & inhibitors
  • Myosins / antagonists & inhibitors
  • Myosins / metabolism*
  • Myosins / physiology
  • Neutrophils / drug effects
  • Neutrophils / metabolism
  • Neutrophils / physiology*
  • Pseudopodia / drug effects
  • Pseudopodia / physiology*
  • Talin / metabolism

Substances

  • Actins
  • Azepines
  • Buffers
  • Enzyme Inhibitors
  • Talin
  • ML 9
  • Myosin-Light-Chain Kinase
  • Myosins
  • Calcium