Stable adhesion and migration of human neutrophils requires phospholipase D-mediated activation of the integrin CD11b/CD18.
The pathways regulating integrin-mediated adhesion during neutrophil migration are incompletely defined. Using a flow-based model in which human neutrophils rolling on P-selectin were activated to migrate by the chemoattractant peptide fMLP, we investigated the role of phospholipase D (PLD). fMLP-stimulated PLD generation of phosphatidate (PtdOH); while inhibition of PtdOH production with butan-1-ol had no effect on the initial immobilisation of rolling neutrophils (supported by activation of constitutively surface-expressed beta(2)-integrin CD11b/CD18) it impaired longer-term stability of adhesion and reduced the rate of migration (supported by activation of de novo-exocytosed CD11b/CD18). PtdOH regulated these processes by controlling activation of exocytosed CD11b/CD18, and appeared to act by directly stimulating phosphatidylinositol 4-phosphate 5-kinase type I to generate phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P(2)). Cell-permeable PtdIns(4,5)P(2) recovered migration of neutrophils after PLD inhibition; PtdIns(4,5)P(2) appeared to act by promoting talin binding to CD18 and hence activating CD11b/CD18, as migration was inhibited when neutrophils were loaded with peptides previously shown to block the interaction between PtdIns(4,5)P(2) and talin or talin and CD18. Thus, these data indicate that PLD-synthesised PtdOH stimulates the generation of PtdIns(4,5)P(2), which in turn mediates talin binding to, and activation of, CD11b/CD18 required for neutrophil stable adhesion and migration.
Powner
Dale J
DJ
CRUK Institute for Cancer Studies, The Medical School, The University of Birmingham, Birmingham, B15 2TT, UK. D.J.Powner.20@bham.ac.uk <D.J.Powner.20@bham.ac.uk>
Pettitt
Trevor R
TR
Anderson
Rhodri
R
Nash
Gerard B
GB
Wakelam
Michael J O
MJ
D039481:CD11b Antigen / Q000378:metabolism*; D018821:CD18 Antigens / Q000378:metabolism*; D002448:Cell Adhesion; D002634:Chemotaxis, Leukocyte; D006801:Humans; D016023:Integrins / Q000378:metabolism; D036904:Leukocyte Rolling; D009504:Neutrophils / Q000166:cytology / Q000502:physiology*; D010712:Phosphatidic Acids / Q000494:pharmacology; D010739:Phospholipase D / Q000502:physiology*
D039481:CD11b Antigen; D018821:CD18 Antigens; C509295:ITGAM protein, human; D016023:Integrins; D010712:Phosphatidic Acids; D010739:Phospholipase D