TRANSFORMING GROWTH-FACTOR-ALPHA AND EPIDERMAL GROWTH-FACTOR ACTIVATEMITOGEN-ACTIVATED PROTEIN-KINASE AND ITS SUBSTRATES IN INTESTINAL EPITHELIAL-CELLS

The signal transduction pathways of mitogenic stimuli in intestinal ep ithelial cells are not clearly understood, We report here a possible s ignaling pathway of two closely related agonists, transforming growth factor-alpha (TGF alpha) and epidermal growth factor (EGF), Both incre ase thymidine incorporation in the intestinal epithelial cell (IEC) li ne IEC-6, This increase is dose dependent and inhibited by the tyrosin e kinase inhibitors genistein and tyrphostin. The addition of either T GF alpha or EGF to IEC-6 cells also stimulates the activities of the t wo forms of mitogen-activated protein kinase, p42(erk2) MAPK and p44(e rk1) MAPK, as evidenced by increased incorporation of radiolabeled pho sphate in myelin basic protein, The main difference between the MAPK a ctivity levels induced by the two agonists is in the intensity of the response. Maximum TGF alpha-induced stimulation of p42(erk2) MAPK acti vity is g-fold at 2 ng/ml, while maximum EGF stimulation is only 4.5-f old at 25 ng/ml. These doses correlated closely with the dose required for maximum thymidine incorporation, The activity of the 90-kDa ribos omal se kinase, a downstream substrate for activated MAPK, is also enh anced as evidenced by increased incorporation of radiolabeled phosphat e in the rsk kinase substrate peptide in IEC-6 cells following stimula tion with either TGF alpha or EGF, This increase correlates closely wi th the stimulus-induced increase in MAPK activity with respect to dose , but the time of increased activity is more prolonged, especially aft er 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 act ivation.