Re-distribution of phospholipase C gamma 2 in macrophage precursors is mediated by the actin cytoskeleton under the control of the Src kinases.
Macrophage colony-stimulating factor (M-CSF) is a growth factor that is known to trigger several signalling pathways through receptor tyrosine kinase activation. We investigated the specific requirements for the activation of phospholipase C gamma 2 (PLC-gamma2) during the differentiation of mouse bone marrow-derived macrophage precursors. M-CSF stimulation induced rapid PLC-gamma2 translocation and phosphorylation from the cytosolic compartment to the cell periphery. Both events were dependent on cytoskeleton integrity and Src kinase activity, but only PLC-gamma2 phosphorylation did not require PI3-kinase activity. Biochemical experiments as well as confocal microscopy analyses indicate that the translocation of PLC-gamma2 is mediated by the direct association of this protein with the actin cytoskeleton. Using GST-fusion proteins containing various deletions of the PLC-gamma2 Src homology region, it was found that PLC-gamma2 binds to F-actin via its SH2 domains, a feature that has equally been found in a co-sedimentation assay. This association, which is increased during actin reorganisation and disrupted by cytoskeleton inhibitors, seems to be a primary means to recruit this enzyme to the cell periphery. These results indicate that, upon M-CSF stimulation, PLC-gamma2 cellular localisation and phosphorylation are strongly dependent on cytoskeleton architecture of the macrophage precursor as well as the PI3-kinase and the Src kinases.
Dearden-Badet
Marie-Thérèse
MT
Centre de Génétique Moléculaire et Cellulaire, UMR CNRS 5534, Bâtiment Gregor Mendel, 16 Rue Raphaël Dubois, 69622 Villeurbanne Cedex, France. dearden@cgmc.univ-lyon1.fr
Mouchiroud
Guy
G
D000199:Actins / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D002454:Cell Differentiation; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D015638:Cytochalasin D / Q000494:pharmacology; D003599:Cytoskeleton / Q000737:chemistry / Q000378:metabolism*; D016173:Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D008264:Macrophages / Q000166:cytology / Q000201:enzymology* / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D009025:Morpholines / Q000494:pharmacology; D015739:Nocodazole / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051966:Phospholipase C gamma / Q000378:metabolism*; D021381:Protein Transport; D011743:Pyrimidines / Q000494:pharmacology; D013997:Time Factors; D019061:src-Family Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*
D000199:Actins; D002867:Chromones; D015638:Cytochalasin D; D016173:Macrophage Colony-Stimulating Factor; D009025:Morpholines; D015739:Nocodazole; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051966:Phospholipase C gamma; D011743:Pyrimidines; D019061:src-Family Kinases; C412373:AG 1879; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one