CHEMOTAXIS OF CHINESE-HAMSTER OVARY CELLS EXPRESSING THE HUMAN NEUTROPHIL FORMYL PEPTIDE RECEPTOR - ROLE OF SIGNAL-TRANSDUCTION MOLECULES AND ALPHA(5)BETA(1) INTEGRIN
Hm. Miettinen et al., CHEMOTAXIS OF CHINESE-HAMSTER OVARY CELLS EXPRESSING THE HUMAN NEUTROPHIL FORMYL PEPTIDE RECEPTOR - ROLE OF SIGNAL-TRANSDUCTION MOLECULES AND ALPHA(5)BETA(1) INTEGRIN, Journal of Cell Science, 111, 1998, pp. 1921-1928
Activation of the N-formyl peptide receptor (FPR) of human neutrophils
by ligands such as N-formylmethionine-leucine-phenylalanine (fMLP) in
duces mobilization of intracellular calcium, cell adhesion, chemotaxis
, superoxide production and degranulation, Chinese hamster ovary (CHO)
cells are normally devoid of FPR and unresponsive to fMLP, but when s
tably transfected with a human FPR cDNA, exhibited some of these same
responses, Specifically, stimulation with fMLP resulted in release of
intracellular calcium and chemotactic migration toward a gradient of f
MLP. As in neutrophils, both processes were inhibited through receptor
desensitization by prior exposure to a higher or equal concentration
of ligand or by treatment with pertussis toxin, Soluble and membrane-b
ound fibronectin greatly increased fMLP-induced chemotaxis of CHO cell
s expressing FPR, but not of wild-type CHO cells, suggesting a role fo
r FPR in activation of integrin function. Evidence for this hypothesis
was obtained by demonstrating that CHO cells expressing FPR rapidly i
ncreased their adhesion to a fibronectin-coated surface after stimulat
ion with fMLP. Both chemotaxis and adhesion were largely inhibited by
RGDS peptide and a function-blocking antibody against as integrin, FPR
-mediated chemotaxis of the CHO transfectants was partly inhibited by
a tyrosine kinase inhibitor, herbimycin A, and blocked by a phosphoino
sitide 3-kinase inhibitor, wortmannin. These data suggest that stimula
tion of CHO FPR transfectants with a gradient of JMLP results in phosp
hoinositide 3-kinase-dependent chemotactic migration, which is enhance
d by binding of activated alpha(5)beta(1) to fibronectin. This non-mye
loid, non-lymphoid fibroblastic cell line will thus serve as a useful
model to investigate additional requirements of signal transduction mo
lecules, adhesion molecules and cytoskeletal elements in FPR-mediated
chemotaxis.