Cholecystokinin stimulates extracellular signal-regulated kinase through activation of the epidermal growth factor receptor, Yes, and protein kinase C. Signal amplification at the level of Raf by activation of protein kinase Cepsilon.
Cholecystokinin (CCK) and related peptides are potent growth factors in the gastrointestinal tract and may be important for human cancer. CCK exerts its growth modulatory effects through G(q)-coupled receptors (CCK(A) and CCK(B)) and activation of extracellular signal-regulated protein kinase 1/2 (ERK1/2). In the present study, we investigated the different mechanisms participating in CCK-induced activation of ERK1/2 in pancreatic AR42J cells expressing both CCK(A) and CCK(B). CCK activated ERK1/2 and Raf-1 to a similar extent as epidermal growth factor (EGF). Inhibition of EGF receptor (EGFR) tyrosine kinase or expression of dominant-negative Ras reduced CCK-induced ERK1/2 activation, indicating participation of the EGFR and Ras in CCK-induced ERK1/2 activation. However, compared with EGF, CCK caused only small increases in tyrosine phosphorylation of the EGFR and Shc, Shc-Grb2 complex formation, and Ras activation. Signal amplification between Ras and Raf in a CCK-induced ERK cascade appears to be mediated by activation of protein kinase Cepsilon (PKCepsilon), because 1) down-modulation of phorbol ester-sensitive PKCs inhibited CCK-induced activation of Ras, Raf, and ERK1/2 without influencing Shc-Grb2 complex formation; 2) PKCepsilon, but not PKCalpha or PKCdelta, was detectable in Raf-1 immunoprecipitates, although CCK activated all three PKC isoenzymes. In addition, the present study provides evidence that the Src family tyrosine kinase Yes is activated by CCK and mediates CCK-induced tyrosine phosphorylation of Shc. Furthermore, we show that CCK-induced activation of the EGFR and Yes is achieved through the CCK(B) receptor. Together, our data show that different signals emanating from the CCK receptors mediate ERK1/2 activation; activation of Yes and the EGFR mediate Shc-Grb2 recruitment, and activation of PKC, most likely PKCepsilon, augments CCK-stimulated ERK1/2 activation at the Ras/Raf level.
Piiper
Albrecht
A
Department of Internal Medicine II, University of the Saarland, D-66421 Homburg/Saar, Germany. inapii@uniklinik-saarland.de
Elez
Robert
R
You
Se-Jong
SJ
Kronenberger
Bernd
B
Loitsch
Stefan
S
Roche
Serge
S
Zeuzem
Stefan
S
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D000818:Animals; D016466:CHO Cells; D002766:Cholecystokinin / Q000494:pharmacology*; D006224:Cricetinae; D015536:Down-Regulation; D004789:Enzyme Activation; D066246:ErbB Receptors / Q000378:metabolism*; D051380:GRB2 Adaptor Protein; D006801:Humans; D015151:Immunoblotting; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D010703:Phorbol Esters / Q000378:metabolism; D010766:Phosphorylation; D011233:Precipitin Tests; D011485:Protein Binding; D011493:Protein Kinase C / Q000378:metabolism*; D051744:Protein Kinase C-epsilon; D021381:Protein Transport; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism*; D051575:Proto-Oncogene Proteins c-yes; D051381:Rats; D029589:Seminal Plasma Proteins / Q000378:metabolism*; D055765:Shc Signaling Adaptor Proteins; D015398:Signal Transduction; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D013997:Time Factors; D015533:Transcriptional Activation; D014162:Transfection; D014443:Tyrosine / Q000378:metabolism; D018631:ras Proteins / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; C495950:Grb2 protein, rat; D010703:Phorbol Esters; D011506:Proteins; C528024:SHC1 protein, human; D029589:Seminal Plasma Proteins; D055765:Shc Signaling Adaptor Proteins; C528035:Shc1 protein, rat; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D014443:Tyrosine; D002766:Cholecystokinin; C497257:Prkce protein, rat; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases; D051575:Proto-Oncogene Proteins c-yes; C489834:YES1 protein, human; D019908:Proto-Oncogene Proteins c-raf; C497255:PRKCE protein, human; D011493:Protein Kinase C; D051744:Protein Kinase C-epsilon; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D018631:ras Proteins