The expression of peptides of the pancreatitis-associated protein/pancreatic stone protein family appears to be increased in response to tissue injury. One member of this group, Reg1α, is expressed in gastric epithelial cells, and may be a gastric growth factor (Fukui et al. 1998). The expression of Reg1α in stomach is controlled by the gastric hormone gastrin (Higham et al. 1999). We have examined the transcriptional mechanisms controlling Reg1α gene expression.
A sequence of 2.1 kb of the promoter region of the rat Reg gene was cloned by PCR and sequenced. Deletional mutants were cloned into a luciferase reporter vector, and luciferase expression was determined in AGS-GR cells (105 cells in 6-well dishes) which express the gastrin/cholecystokinin-B receptor. Gastrin (1 nM, 8 h) stimulated luciferase expression of the smallest deletional mutant (104 bp upstream of the transcriptional start site) 5.2 ± 0.4-fold (mean ± S.E.M., n = 12); there was no significant difference in the relative increase in luciferase activity of the 104 bp and the 2.1 kb promoter sequences in response to gastrin. Mutations within the 104 bp sequence suggested a putative Sp1 site was important for basal and gastrin-stimulated expression. The effect of gastrin on the 104 bp sequence was almost abolished by Clostridium botulinum C3-exoenzyme (24 h, 800 ng ml-1), which inhibits the small GTPase, RhoA. Consistent with this observation, when cells were co-transfected with a dominant negative RhoA construct (N19RhoA; 0.5 mg) and the 104 bp Reg-luciferase expression vector (0.5 mg), the response to gastrin was significantly inhibited (1.6 ± 0.2-fold, P < 0.05, unpaired t test). Moreover, co-transfection of cells with a constitutively active form of RhoA (L63RhoA) significantly increased expression of the Reg-luciferase vector (5.7 ± 0.4-fold, P < 0.05); in contrast, constitutively active forms of the related GTPases, Rac1 and cdc42 had little or no stimulatory effect on Reg1α expression.
The data suggest that expression of the Reg gene is regulated by gastrin and that activation of RhoA is part of the transduction mechanism.