Regulation of intestinal uroguanylin/guanylin receptor-mediated responses by mucosal acidity

Abstract

Guanylin and uroguanylin are intestinal peptides that stimulate chloride secretion by activating a common set of receptor-guanylate cyclase signaling molecules located on the mucosal surface of enterocytes. High mucosal acidity, similar to the pH occurring within the f luid microclimate domain at the mucosal surface of the intestine, markedly enhances the cGMP accumulation responses of T84 human intestinal cells to uroguanylin. In contrast, a mucosal acidity of pH 5.0 renders guanylin essentially inactive. T84 cells were used as a model epithelium to further explore the concept that mucosal acidity imposes agonist selectivity for activation of the intestinal receptors for uroguanylin and guanylin, thus providing a rationale for the evolution of these related peptides. At an acidic mucosal pH of 5.0, uroguanylin is 100-fold more potent than guanylin, but at an alkaline pH of 8.0 guanylin is more potent than uroguanylin in stimulating intracellular cGMP accumulation and transepithelial chloride secretion. The relative affinities of uroguanylin and guanylin for binding to receptors on the mucosal surface of T84 cells is inf luenced dramatically by mucosal acidity, which explains the strong pH dependency of the cGMP and chloride secretion responses to these peptides. The guanylin-binding affinities for peptide-receptor interaction were reduced by 100-fold at pH 5 versus pH 8, whereas the affinities of uroguanylin for these receptors were increased 10-fold by acidic pH conditions. Deletion of the N-terminal acidic amino acids in uroguanylin demonstrated that these residues are responsible for the increase in binding affinities that are observed for uroguanylin at acidic pH. We conclude that guanylin and uroguanylin evolved distinctly different structures, which enables both peptides to regulate, in a pHdependent fashion, the activity of receptors that control intestinal salt and water transport via cGMP.Guanylin and uroguanylin are structurally related peptides that were isolated from intestinal mucosa and urine (1-5). A receptor for guanylin and uroguanylin that has been identified at the molecular level is a transmembrane form of guanylate cyclase, termed GC-C (6). This membrane protein was originally discovered as an intestinal receptor for the heat-stable toxin (ST) peptides, which are secreted intraluminally by enteric bacteria that cause traveler's diarrhea (7). Bacterial ST peptides are related in primary structure to uroguanylin and guanylin, thus acting as molecular mimics of the enteric peptide hormones (reviewed in refs. 8 and 9). Membrane receptor-guanylate cyclases are found on the luminal surface of enterocytes throughout the small and large intestine and in other epithelia (10-13). Binding of peptide agonists to an extracellular domain of the receptor activates the intracellular catalytic domain producing the second messenger cGMP within target enterocytes (1-6). Intracellular cGMP stimulates transepithelial chloride secretion by regulating the phosphorylation state and chloride channel activit...

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“…These peptides share sequence homology, have a tertiary structure stabilized by intrachain disulfide bonds, and exert their (patho)physiological effects by binding to GC-C and inducing cGMP accumulation. Uroguanylin, which is highly expressed in stomach, duodenum, and jejunum, is 100-fold more potent than guanylin at acidic pH (18,19). In contrast, guanylin is more abundant in ileum and colon and is 4-fold more potent than uroguanylin at a pH of 8.0 (18,19).…”

“…One hypothesis suggests a role for these peptides in the paracrine and autocrine regulation of f luid and electrolyte homeostasis in the intestine. Also, these peptides are expressed in extra-intestinal sites, including the kidney, suggesting that they may comprise one limb of an endocrine feedback loop that integrates the intestine into mechanisms regulating volume homeostasis (18,19). This function for GC-C is analogous to that for GC-A and GC-B and their agonists, the natriuretic peptides, which regulate f luid and electrolyte secretion in the kidney and play a central role in volume homeostasis (11).…”

“…Uroguanylin, which is highly expressed in stomach, duodenum, and jejunum, is 100-fold more potent than guanylin at acidic pH (18,19). In contrast, guanylin is more abundant in ileum and colon and is 4-fold more potent than uroguanylin at a pH of 8.0 (18,19). These endogenous GC-C agonists may regulate physiological processes in distinct regions of the intestine, modulated by local pH.…”

A novel role for uroguanylin in the regulation of sodium balance

“…Studies using the T84 colon cell model may have provided some insights into some molecular features of the uroguanylin molecule that influence its biological activity. For example, the potency of uroguanylin remains high when the growth medium pH is reduced to an acidic condition, which in turn greatly reduces the potency of guanylin for activation of chloride secretion in T84 cells (18). The effects of pH appear to be manifest in the affinity of these peptides for interaction with R-GC-C.…”

Cellular Effects of Guanylin and Uroguanylin

“…Conversely, GN is more potent at pH 8 compared with pH 5 ( Figure 4). As this pH dependence affects the ligand/receptor interaction, it was suggested that the Nterminal ends of the UGN and GN molecules are responsible for this pH dependence (50). The duodenum secretes bicarbonate to neutralize the acidic pH of the stomach contents that enter the duodenum.…”