Role of metalloproteinase-dependent EGF receptor activation in alpha-adrenoceptor-stimulated MAP kinase phosphorylation in GT1-7 neurons.
Adrenoceptors (ARs) are involved in the regulation of gonadotropin-releasing hormone (GnRH) release from native and immortalized hypothalamic (GT1-7) neurons. However, the AR-mediated signaling mechanisms and their functional significance in these cells are not known. Stimulation of GT1-7 cells with the alpha1-AR agonist, phenylephrine (Phe), causes phosphorylation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) mitogen-activated protein (MAP) kinases that is mediated by protein kinase C (PKC)-dependent transactivation of the epidermal growth factor receptor (EGF-R). Phe stimulation causes shedding of the soluble ligand, heparin-binding EGF (HB-EGF), as a consequence of matrix metalloproteinase (MMP) activation. Phe-induced phosphorylation of the EGF-R, and subsequently of Shc and ERK1/2, was attenuated by inhibition of MMP or HB-EGF with the selective inhibitor, CRM197, or by a neutralizing antibody. In contrast, phosphorylation of the EGF-R, Shc and ERK1/2 by EGF and HB-EGF was independent of PKC and MMP activity. Moreover, inhibition of Src attenuated ERK1/2 responses by Phe, but not by HB-EGF and EGF, indicating that Src acts upstream of the EGF-R. Consistent with a potential role of reactive oxygen species (ROS), Phe-induced phosphorylation of EGF-R was attenuated by the antioxidant, N-acetylcysteine. These data suggest that activation of the alpha1-AR causes phosphorylation of ERK1/2 through activation of PKC, ROS and Src, and shedding of HB-EGF, which binds to and activates the EGF-R.
Shah
Bukhtiar H
BH
Endocrinology and Reproduction Research Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892-4510, USA. shahb@mail.nih.gov
Shah
Farzana B
FB
Catt
Kevin J
KJ
D000316:Adrenergic alpha-Agonists / Q000494:pharmacology; D000818:Animals; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D045726:Metalloproteases / Q000378:metabolism*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D010656:Phenylephrine / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000502:physiology; D051057:Proto-Oncogene Proteins c-akt / Q000502:physiology; D018341:Receptors, Adrenergic, alpha-2 / Q000502:physiology*; D015533:Transcriptional Activation / Q000502:physiology; D019061:src-Family Kinases / Q000502:physiology
D000316:Adrenergic alpha-Agonists; D018341:Receptors, Adrenergic, alpha-2; D010656:Phenylephrine; D066246:ErbB Receptors; D019061:src-Family Kinases; D051057:Proto-Oncogene Proteins c-akt; D011493:Protein Kinase C; D048049:Extracellular Signal-Regulated MAP Kinases; D020928:Mitogen-Activated Protein Kinases; D045726:Metalloproteases