MOLECULAR MECHANISMS FOR THE GROWTH-FACTOR ACTION OF GASTRIN

We have previously observed that gastrin has a cholecystokinin B (CCK- B) receptor-mediated growth-promoting effect on the AR42J rat pancreat ic acinar cell Line and that this effect is paralleled by induction of expression of the early response gene c-fas. We undertook these exper iments to elucidate the mechanism for induction of c-fos and the linka ge of this action to the trophic effects of gastrin. Gastrin (0.1-10 n M) dose dependently induced luciferase activity in AR42J cells transfe cted with a construct consisting of a luciferase reporter gene coupled to the serum response element (SRE) of the c-fos promoter. This effec t was blocked by the specific CCK-B receptor antagonist D2 but not by the specific CCK-A receptor antagonist L-364,718 or by pertussis toxin , indicating that gastrin targets the SRE via specific CCK-B receptors through a mechanism independent of Gi. Inhibition of protein kinase C (PKC) either by prolonged (24 h) exposure of the cells to the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (100 nM) or by incubation with the selective inhibitor GF-109203X (3.5 mu M) resulted in an 80% reduction in luciferase activity. Similar results were observed in the presence of the specific extracellular signal-regulated kinase (ERK) kinase (MEK) inhibitor PD-98059 (50 mu M). We measured ERK2 activity i n AR42J cells via in-gel kinase assays and observed that gastrin (1 pM -100 nM) induced ERK2 enzyme activity in a dose-dependent manner. Addi tion of GF-109203X and PD-98059, either alone or in combination, produ ced, respectively, partial and total inhibit ion of gastrin-induced ER K2 activity. Gastrin induction of ERK2 activity also resulted in a thr eefold increase in the transcriptional activity of Elk-1, a factor kno wn to bind to the c-fos SRE and to be phosphorylated and activated by ERK2. PD-98059 blocked the growth-promoting effect of gastrin on the A R42J cells, demonstrating that this effect depends on activation of ME K. Our data lead us to conclude that the trophic actions of gastrin ar e mediated by ERK2-induced c-fos gene expression via PKC-dependent and -independent pathways.