Transforming growth factor-alpha and epidermal growth factor activate mitogen-activated protein kinase and its substrates in intestinal epithelial cells.
The signal transduction pathways of mitogenic stimuli in intestinal epithelial cells are not clearly understood. We report here a possible signaling pathway of two closely related agonists, transforming growth factor-alpha (TGF alpha) and epidermal growth factor (EGF). Both increase thymidine incorporation in the intestinal epithelial cell (IEC) line IEC-6. This increase is dose dependent and inhibited by the tyrosine kinase inhibitors genistein and tyrphostin. The addition of either TGF alpha or EGF to IEC-6 cells also stimulates the activities of the two forms of mitogen-activated protein kinase, p42erk2 MAPK and p44erk1 MAPK, as evidenced by increased incorporation of radiolabeled phosphate in myelin basic protein. The main difference between the MAPK activity levels induced by the two agonists is in the intensity of the response. Maximum TGF alpha-induced stimulation of p42erk2 MAPK activity is 9-fold at 2 ng/ml, while maximum EGF stimulation is only 4.5-fold at 25 ng/ml. These doses correlated closely with the dose required for maximum thymidine incorporation. The activity of the 90-kDa ribosomal S6 kinase, a downstream substrate for activated MAPK, is also enhanced as evidenced by increased incorporation of radiolabeled phosphate in the rsk kinase substrate peptide in IEC-6 cells following stimulation with either TGF alpha or EGF. This increase correlates closely with the stimulus-induced increase in MAPK activity with respect to dose, but the time of increased activity is more prolonged, especially after EGF stimulation. TGF alpha induced the synthesis of both c-Fos and c-Myc, two nuclear substrates for MAPK, and increased c-fos and c-myc message levels as well. However, c-Jun protein and c-jun mRNA were not induced. The increase in IEC-6 cell proliferation in response to TGF alpha and EGF stimulation may then be due, in part, to an increase in immediate early gene expression as a direct result of MAPK and RSK activation.
Oliver
B L
BL
Department of Physiology, University of Connecticut Health Center, Farmington 06030, USA.
Sha'afi
R I
RI
Hajjar
J J
JJ
D000818:Animals; D015152:Blotting, Northern; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002396:Catechols / Q000494:pharmacology; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D004247:DNA / Q000096:biosynthesis; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D004847:Epithelial Cells; D004848:Epithelium / Q000187:drug effects / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D016762:Genes, fos / Q000187:drug effects; D016758:Genes, jun / Q000187:drug effects; D016259:Genes, myc / Q000187:drug effects; D019833:Genistein; D007413:Intestinal Mucosa / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D007421:Intestine, Small / Q000378:metabolism; D007529:Isoflavones / Q000494:pharmacology; D007700:Kinetics; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D004676:Myelin Basic Protein / Q000378:metabolism; D009570:Nitriles / Q000494:pharmacology; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D016760:Proto-Oncogene Proteins c-fos / Q000096:biosynthesis; D016755:Proto-Oncogene Proteins c-jun / Q000096:biosynthesis; D016271:Proto-Oncogene Proteins c-myc / Q000096:biosynthesis; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis; D051381:Rats; D013379:Substrate Specificity; D013936:Thymidine / Q000378:metabolism; D016211:Transforming Growth Factor alpha / Q000494:pharmacology*; D020032:Tyrphostins
D002396:Catechols; D004791:Enzyme Inhibitors; D007529:Isoflavones; D004676:Myelin Basic Protein; D009570:Nitriles; D016760:Proto-Oncogene Proteins c-fos; D016755:Proto-Oncogene Proteins c-jun; D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger; D016211:Transforming Growth Factor alpha; D020032:Tyrphostins; D004815:Epidermal Growth Factor; D004247:DNA; D019833:Genistein; D011505:Protein-Tyrosine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; C060641:tyrphostin A23; D013936:Thymidine