TY - JOUR
T1 - Erythropoietin receptor Y479 couples to ERK1/2 activation via recruitment of phospholipase Cγ
AU - Halupa, Adrienne
AU - Chohan, Manprit
AU - Stickle, Natalie H.
AU - Beattie, Bryan K.
AU - Miller, Barbara A.
AU - Barber, Dwayne L.
N1 - Funding Information:
This research was supported by a grant from Canadian Institutes of Health Research. AH was supported by a Premier's Research Excellence Award. DLB is a National Cancer Institute of Canada Research Scientist. We thank Stanley Liu and Jane McGlade for assistance in COLT screening. We acknowledge the generous gifts of reagents from Graham Carpenter, Vanderbilt University, Nashville, TN and Steve Watson, University of Oxford, UK. We would like to thank Jane McGlade, Sam Benchimol, Sean Egan, Eleanor Fish, and Mark Koury and members of the Barber laboratory for helpful comments on the manuscript.
PY - 2005/9/10
Y1 - 2005/9/10
N2 - Red blood cell development is primarily controlled by erythropoietin (EPO). Several studies have revealed the importance of EPO-R Y343 and Y479 for erythroid cell growth, differentiation, and survival. In order to isolate critical signaling proteins that bind to EPO-R, we initiated a Cloning of Ligand Target (COLT) screen using a murine embryonic day 16 phage library and a biotinylated EPO-R Y343 phosphopeptide. One of the clones isolated encodes Phospholipase C (PLC)γ1. PLCγ1 is rapidly tyrosine phosphorylated upon EPO stimulation and associates with EPO-R in an SH2-domain-dependent manner. Although PLCγ1 bound EPO-R Y343, Y401, Y429, Y431, and Y479 in the COLT screen, PLCγ1 required Y479 for association with EPO-R in Ba/F3-EPO-R cells. Studies have identified EPO-R Y479 as important for ERK activation. Since PI3-kinase binds EPO-R Y479, one group has suggested that ERK activation downstream of PI3-kinase accounts for the importance of this residue in EPO signaling. However, we show that inhibition of PI3-kinase does not abolish ERK activation. Furthermore, we demonstrate interaction of PLCγ1 with Grb2 and SOS2. Hence, we have identified a novel adapter function for PLCγ1 in EPO signaling in which recruitment of PLCγ1 to EPO-R may lead to activation of the ERK pathway.
AB - Red blood cell development is primarily controlled by erythropoietin (EPO). Several studies have revealed the importance of EPO-R Y343 and Y479 for erythroid cell growth, differentiation, and survival. In order to isolate critical signaling proteins that bind to EPO-R, we initiated a Cloning of Ligand Target (COLT) screen using a murine embryonic day 16 phage library and a biotinylated EPO-R Y343 phosphopeptide. One of the clones isolated encodes Phospholipase C (PLC)γ1. PLCγ1 is rapidly tyrosine phosphorylated upon EPO stimulation and associates with EPO-R in an SH2-domain-dependent manner. Although PLCγ1 bound EPO-R Y343, Y401, Y429, Y431, and Y479 in the COLT screen, PLCγ1 required Y479 for association with EPO-R in Ba/F3-EPO-R cells. Studies have identified EPO-R Y479 as important for ERK activation. Since PI3-kinase binds EPO-R Y479, one group has suggested that ERK activation downstream of PI3-kinase accounts for the importance of this residue in EPO signaling. However, we show that inhibition of PI3-kinase does not abolish ERK activation. Furthermore, we demonstrate interaction of PLCγ1 with Grb2 and SOS2. Hence, we have identified a novel adapter function for PLCγ1 in EPO signaling in which recruitment of PLCγ1 to EPO-R may lead to activation of the ERK pathway.
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U2 - 10.1016/j.yexcr.2005.04.030
DO - 10.1016/j.yexcr.2005.04.030
M3 - Article
C2 - 15953601
AN - SCOPUS:23944506056
VL - 309
SP - 1
EP - 11
JO - Experimental Cell Research
JF - Experimental Cell Research
SN - 0014-4827
IS - 1
ER -