P2Y2 nucleotide receptors mediate metalloprotease-dependent phosphorylation of epidermal growth factor receptor and ErbB3 in human salivary gland cells.
The G protein-coupled receptor P2Y(2) nucleotide receptor (P2Y(2)R) has been shown to be up-regulated in a variety of tissues in response to stress or injury. Recent studies have suggested that P2Y(2)Rs may play a role in immune responses, wound healing, and tissue regeneration via their ability to activate multiple signaling pathways, including activation of growth factor receptors. Here, we demonstrate that in human salivary gland (HSG) cells, activation of the P2Y(2)R by its agonist induces phosphorylation of ERK1/2 via two distinct mechanisms, a rapid, protein kinase C-dependent pathway and a slower and prolonged, epidermal growth factor receptor (EGFR)-dependent pathway. The EGFR-dependent stimulation of UTP-induced ERK1/2 phosphorylation in HSG cells is inhibited by the adamalysin inhibitor tumor necrosis factor-alpha protease inhibitor or by small interfering RNA that selectively silences ADAM10 and ADAM17 expression, suggesting that ADAM metalloproteases are required for P2Y(2)R-mediated activation of the EGFR. G protein-coupled receptors have been shown to promote proteolytic release of EGFR ligands; however, neutralizing antibodies to known ligands of the EGFR did not inhibit UTP-induced EGFR phosphorylation. Immunoprecipitation experiments indicated that UTP causes association of the EGFR with another member of the EGF receptor family, ErbB3. Furthermore, stimulation of HSG cells with UTP induced phosphorylation of ErbB3, and silencing of ErbB3 expression inhibited UTP-induced phosphorylation of both ErbB3 and EGFR. UTP-induced phosphorylation of ErbB3 and EGFR was also inhibited by silencing the expression of the ErbB3 ligand neuregulin 1 (NRG1). These results suggest that P2Y(2)R activation in salivary gland cells promotes the formation of EGFR/ErbB3 heterodimers and metalloprotease-dependent neuregulin 1 release, resulting in the activation of both EGFR and ErbB3.
Ratchford
Ann M
AM
Department of Biochemistry, University of Missouri, Christopher S. Bond Life Sciences Center, Columbia, Missouri 65211, USA.
Baker
Olga J
OJ
Camden
Jean M
JM
Rikka
Shivaji
S
Petris
Michael J
MJ
Seye
Cheikh I
CI
Erb
Laurie
L
Weisman
Gary A
GA
D051722:ADAM Proteins / Q000235:genetics / Q000378:metabolism; D000072197:ADAM10 Protein; D000072198:ADAM17 Protein; D053829:Amyloid Precursor Protein Secretases / Q000235:genetics / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000378:metabolism; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D045726:Metalloproteases / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D020890:Neuregulin-1 / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D055503:Protein Multimerization; D058910:Purinergic P2 Receptor Agonists; D011799:Quinazolines; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D020893:Receptor, ErbB-3 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018048:Receptors, Purinergic P2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D058473:Receptors, Purinergic P2Y2; D012469:Salivary Glands / Q000166:cytology* / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D020032:Tyrphostins / Q000378:metabolism
D004791:Enzyme Inhibitors; D008565:Membrane Proteins; D020890:Neuregulin-1; C546385:P2RY2 protein, human; C546386:P2ry2 protein, mouse; D058910:Purinergic P2 Receptor Agonists; D011799:Quinazolines; D034741:RNA, Small Interfering; D018048:Receptors, Purinergic P2; D058473:Receptors, Purinergic P2Y2; D020032:Tyrphostins; C101044:RTKI cpd; D066246:ErbB Receptors; D020893:Receptor, ErbB-3; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D053829:Amyloid Precursor Protein Secretases; D045726:Metalloproteases; D051722:ADAM Proteins; D000072197:ADAM10 Protein; C491236:ADAM10 protein, human; D000072198:ADAM17 Protein; C000606462:ADAM17 protein, human; C000606463:Adam17 protein, mouse