Phospholipase C gamma complexes with ligand-activated platelet-derived growth factor receptors. An intermediate implicated in phospholipase activation.
Several steps implicated in platelet-derived growth factor (PDGF) receptor-coupled signaling are activated by PDGF exposure at 0-4 degrees C. These include receptor self-phosphorylation, physical association with and phosphorylation of phospholipase C gamma (PLC gamma). Reduced temperature blocks PDGF internalization, making it possible to dissociate bound PDGF after PLC gamma tyrosine phosphorylation. We addressed the functional consequences of PDGF dissociation from intact cell PDGF receptors. PDGF exposure at 0-4 degrees C for 15 min stimulated self-phosphorylation of a subpopulation of BALB/c 3T3 cell PDGF beta-type receptors (35%) and initiated subsequent inositol phosphate production. A small fraction of cellular PLC gamma (1-3%) coprecipitated with ligand-activated PDGF receptors; 3-5% of cellular PLC gamma acquired phosphotyrosine. The PLC gamma coprecipitating with PDGF receptors did not contain detectible phosphotyrosine. Phosphotyrosine antibody recovered similar amounts of PLC gamma from soluble and particulate fractions of PDGF-stimulated cells. Acid dissociation of bound PDGF from receptor caused rapid dephosphorylation of PDGF receptors and PCL gamma, and interrupted PLC gamma-PDGF receptor coprecipitation. Orthovanadate blocked tyrosine dephosphorylation of both PDGF receptors and PLC gamma and stabilized coprecipitation. Orthovanadate reversed the acid wash effect to abrogate PDGF-stimulated inositol phosphate production. PDGF receptor remains competent to coprecipitate with PLC gamma and stimulate PLC-mediated inositol phosphate production if PDGF-induced receptor phosphorylation is maintained. Formation of a coprecipitable PDGF receptor-PLC gamma complex appears required for PDGF-stimulated inositol phosphate production.
Kumjian
D A
DA
Department of Medicine, Vanderbilt University, Nashville, Tennessee 37232.
Barnstein
A
A
Rhee
S G
SG
Daniel
T O
TO
D015153:Blotting, Western; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D006863:Hydrogen-Ion Concentration; D007295:Inositol Phosphates / Q000096:biosynthesis; D008024:Ligands; D010750:Phosphoproteins / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism*; D017479:Receptors, Platelet-Derived Growth Factor; D010738:Type C Phospholipases / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism; D014638:Vanadates / Q000494:pharmacology
D007295:Inositol Phosphates; D008024:Ligands; D010750:Phosphoproteins; D011956:Receptors, Cell Surface; D014638:Vanadates; D014443:Tyrosine; D017479:Receptors, Platelet-Derived Growth Factor; D010738:Type C Phospholipases