Rap2B-dependent stimulation of phospholipase C-epsilon by epidermal growth factor receptor mediated by c-Src phosphorylation of RasGRP3.
Receptor tyrosine kinase regulation of phospholipase C-epsilon (PLC-epsilon), which is under the control of Ras-like and Rho GTPases, was studied with HEK-293 cells endogenously expressing PLC-coupled epidermal growth factor (EGF) receptors. PLC and Ca(2+) signaling by the EGF receptor, which activated both PLC-gamma1 and PLC-epsilon, was specifically suppressed by inactivation of Ras-related GTPases with clostridial toxins and expression of dominant-negative Rap2B. EGF induced rapid and sustained GTP loading of Rap2B, binding of Rap2B to PLC-epsilon, and Rap2B-dependent translocation of PLC-epsilon to the plasma membrane. GTP loading of Rap2B by EGF was inhibited by chelation of intracellular Ca(2+) and expression of lipase-inactive PLC-gamma1 but not of PLC-epsilon. Expression of RasGRP3, a Ca(2+)/diacylglycerol-regulated guanine nucleotide exchange factor for Ras-like GTPases, but not expression of various other exchange factors enhanced GTP loading of Rap2B and PLC/Ca(2+) signaling by the EGF receptor. EGF induced tyrosine phosphorylation of RasGRP3, but not RasGRP1, apparently caused by c-Src; inhibition of c-Src interfered with EGF-induced Rap2B activation and PLC stimulation. Collectively, these data suggest that the EGF receptor triggers activation of Rap2B via PLC-gamma1 activation and tyrosine phosphorylation of RasGRP3 by c-Src, finally resulting in stimulation of PLC-epsilon.
Stope
Matthias B
MB
Institut für Pharmakologie, Universitätsklinikum Essen, Hufelandstrasse 55, D-45122 Essen, Germany. martina.schmidt@uni-essen.de
Vom Dorp
Frank
F
Szatkowski
Daniel
D
Böhm
Anja
A
Keiper
Melanie
M
Nolte
Jan
J
Oude Weernink
Paschal A
PA
Rosskopf
Dieter
D
Evellin
Sandrine
S
Jakobs
Karl H
KH
Schmidt
Martina
M
D000081247:CSK Tyrosine-Protein Kinase; D066246:ErbB Receptors / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism*; D006801:Humans; D054801:Phosphoinositide Phospholipase C; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D013997:Time Factors; D010738:Type C Phospholipases / Q000378:metabolism*; D020668:rap GTP-Binding Proteins / Q000378:metabolism*; D020663:ras Guanine Nucleotide Exchange Factors; D019061:src-Family Kinases
D020662:Guanine Nucleotide Exchange Factors; C413212:RASGRP3 protein, human; D020663:ras Guanine Nucleotide Exchange Factors; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; D019061:src-Family Kinases; C000631346:CSK protein, human; D010738:Type C Phospholipases; D054801:Phosphoinositide Phospholipase C; C423320:phospholipase C epsilon; C065562:RAP2B protein, human; D020668:rap GTP-Binding Proteins