Effect of Carboxypeptidase E Deficiency on Progastrin Processing and Gastrin Messenger Ribonucleic Acid Expression in Mice with the fat Mutation*

Proforms of gastrointestinal peptides are cleaved at paired basic residues into intermediate forms. Paired basic residues at the Cterminal then are excised by carboxypeptidases before the peptide is amidated. An obese mouse, called Cpe/Cpe, has a missense mutation in carboxypeptidase E (CPE) with no pancreatic CPE activity and a reduced processing of pancreatic proinsulin to insulin. The purpose of this study was 1) to look for the presence of CPE in the antrum of the stomach, duodenum, and colon in the Cpe/Cpe mouse; 2) to determine whether CPE is involved in the processing of progastrin (Pro-G) to its carboxyl-terminal amidated form; and 3) to determine whether a decrease in amidated gastrin results in an upregulation of stomach gastrin messenger RNA (mRNA) levels. In Cpe/Cpe mice, CPE activity was absent in the antrum and colon. In Cpe/Cpe mice, amidated gastrin levels were reduced significantly. Levels of the precursor for amidated gastrin (gastrin-Gly-ArgArg) were markedly elevated. Gastrin mRNA levels were increased approximately 2-fold over the levels in Cpe/Cpe mice. These results indicate that CPE is needed for processing progastrin to gastrin in the stomach and that amidated gastrin exerts an inhibitory feedback effect on gastrin mRNA levels. (Endocrinology 138: 1959–1963, 1997) G IS an endocrine peptide localized primarily in the antral G cells of the stomach mucosa (1). One of the major functions of gastrin is the regulation of gastric acid secretion by the parietal cells of the stomach (1). Gastrin also has trophic effects on the gastrointestinal tract and pancreas (2–6); it affects stomach muscle contraction (7), and it stimulates fractional sodium excretion by the kidney while increasing renal plasma flow (8). Gastrin is processed posttranslationally to intermediate forms, gastrin-Gly-Arg-Arg and gastrin-Gly (9). Although amidation of the COOH-terminal of gastrin is thought to be essential for biological activity (10), there is recent evidence to show that gastrin-Gly can stimulate acid secretion (11) and can have a trophic action (12, 13). In addition, although there is no evidence showing that gastrin-Gly-Arg-Arg itself is active biologically, a recent report indicates that elevated levels of progastrin can increase the bromodeoxyuridine labeling index of the colon (14). An obese mouse model (Cpe/Cpe) has been described that has hyperproinsulinemia and late-onset obesity (15). These Cpe/Cpe mice have a missense mutation in carboxypeptidase E (CPE), with marginal pancreatic CPE-like activity and a reduced capacity to process pancreatic proinsulin to insulin. The missense mutation converts Ser into Pro, a change that eliminates CPE activity (16). CPE is a processing enzyme involved in the excision of paired basic residues remaining at the carboxyl-terminus after cleavage by a prohormone convertase of many endocrine and neuroendocrine peptides (17). Whether CPE is involved in processing progastrin to biologically active amidated gastrin is not known. A deficiency of CPE activity in the stomach may result in the excessive production of intermediate gastrin peptides with C-terminal dibasic residue extensions. In addition, it is conceivable that gastrin messenger RNA (mRNA) expression in the stomach is simultaneously increased as a result of the reduced processing of progastrin to biologically active COOH terminal-amidated gastrin and lower systemic circulatory levels of amidated gastrin. In the present study, we examined the processing of progastrin to gastrin in the antrum of the stomach, gastrointestinal CPE activity, and antral gastrin mRNA levels of Cpe/Cpe mice. The specific detection of amidated gastrin and its proforms was facilitated in part by the availability of specific and sensitive antisera for amidated gastrin and gastrin-Gly-Arg-Arg (18, 19). Materials and Methods