Carbachol stimulates transactivation of epidermal growth factor receptor and mitogen-activated protein kinase in T84 cells. Implications for carbachol-stimulated chloride secretion
| Autor: | S J, Keely, J M, Uribe, K E, Barrett |
|---|---|
| Rok vydání: | 1998 |
| Předmět: |
Receptor
Muscarinic M3 Transcriptional Activation Colon Proteins Biological Transport Tyrphostins Receptors Muscarinic ErbB Receptors Adaptor Proteins Vesicular Transport Chlorides Shc Signaling Adaptor Proteins Calcium-Calmodulin-Dependent Protein Kinases Quinazolines Carbachol Intestinal Mucosa Phosphorylation Protein Kinase C Adaptor Proteins Signal Transducing GRB2 Adaptor Protein Protein Binding Signal Transduction |
| Zdroj: | The Journal of biological chemistry. 273(42) |
| ISSN: | 0021-9258 |
| Popis: | We have examined the role of tyrosine phosphorylation in regulation of calcium-dependent chloride secretion across T84 colonic epithelial cells. The calcium-mediated agonist carbachol (CCh, 100 microM) stimulated a time-dependent increase in tyrosine phosphorylation of a range of proteins (with molecular masses ranging up to 180 kDa) in T84 cells. The tyrosine kinase inhibitor, genistein (5 microM), significantly potentiated chloride secretory responses to CCh, indicating a role for CCh-stimulated tyrosine phosphorylation in negative regulation of CCh-stimulated secretory responses. Further studies revealed that CCh stimulated an increase in both phosphorylation and activity of the extracellular signal-regulated kinase (ERK) isoforms of mitogen-activated protein kinase. Chloride secretory responses to CCh were also potentiated by the mitogen-activated protein kinase inhibitor, PD98059 (20 microM). Phosphorylation of ERK in response to CCh was mimicked by the protein kinase C (PKC) activator, phorbol myristate acetate (100 nM), but was not altered by the PKC inhibitor GF 109203X (1 microM). ERK phosphorylation was also induced by epidermal growth factor (EGF) (100 ng/ml). Immunoprecipitation/Western blot studies revealed that CCh stimulated tyrosine phosphorylation of the EGF receptor (EGFr) and increased co-immunoprecipitation of the adapter proteins, Shc and Grb2, with the EGFr. An inhibitor of EGFr phosphorylation, tyrphostin AG1478 (1 microM), reversed CCh-stimulated phosphorylation of both EGFr and ERK. Tyrphostin AG1478 also potentiated chloride secretory responses to CCh. We conclude that CCh activates ERK in T84 cells via a mechanism involving transactivation of the EGFr, and that this pathway constitutes an inhibitory signaling pathway by which chloride secretory responses to CCh may be negatively regulated. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|