Phosphatidylinositol 3-kinase mediates the inhibitory effect of epidermal growth factor on calcium-dependent chloride secretion.
Epidermal growth factor (EGF) and carbachol both inhibit calcium-activated chloride secretion by the human colonic epithelial cell line, T84. Although the inhibitory mechanism for the carbachol effect involves the 3,4,5,6-isomer of inositol tetrakisphosphate, the mechanisms responsible for the EGF effect have not yet been fully elucidated. Here, we studied the role of phosphatidylinositol 3-kinase (PI 3-kinase) in the inhibitory effect of EGF. The PI 3-kinase inhibitor, wortmannin, slightly increased basal chloride secretion and potentiated the secretory response to thapsigargin. Wortmannin also partially reversed EGF-induced, but not carbachol-induced, inhibition of thapsigargin-stimulated chloride secretion. Wortmannin alone had no effect on carbachol- or histamine-induced chloride secretion and completely reversed EGF-induced inhibition of the secretory response to these agonists. EGF, carbachol, histamine, and thapsigargin all increased levels of the 85-kDa regulatory subunit of PI 3-kinase in antiphosphotyrosine immunoprecipitates. However, only EGF significantly increased levels of the 110-kDa catalytic subunit. Furthermore, only EGF increased PI 3-kinase activity in an in vitro kinase assay. High levels of phosphatidylinositol (3)-monophosphate were present in unstimulated cells and significantly reduced by wortmannin. EGF, but not carbachol, rapidly increased levels of phosphatidylinositol (3,4)-bisphosphate and phosphatidylinositol (3,4,5)-trisphosphate. Production of these lipids was also sensitive to wortmannin. Our data suggest that EGF activates PI 3-kinase and that its lipid products may mediate the inhibitory effect of EGF on calcium-dependent chloride secretion. Our data also suggest that a phosphatidylinositol-specific 3-kinase activity is present in unstimulated T84 cells and may regulate production of phosphatidylinositol (3)-monophosphate and basal secretory tone.
Uribe
J M
JM
Department of Medicine, University of California, San Diego, School of Medicine, San Diego, California 92103, USA.
Keely
S J
SJ
Traynor-Kaplan
A E
AE
Barrett
K E
KE
D000730:Androstadienes / Q000494:pharmacology; D002118:Calcium / Q000037:antagonists & inhibitors* / Q000378:metabolism; D002217:Carbachol / Q000494:pharmacology; D002384:Catalysis; D002460:Cell Line; D002712:Chlorides / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D006801:Humans; D008055:Lipids / Q000096:biosynthesis; D019869:Phosphatidylinositol 3-Kinases; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000077191:Wortmannin
D000730:Androstadienes; D002712:Chlorides; D004791:Enzyme Inhibitors; D008055:Lipids; D004815:Epidermal Growth Factor; D002217:Carbachol; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D002118:Calcium; D000077191:Wortmannin