INACTIVATION OF THE MOUSE GUANYLIN GENE AND ITS REGULATION DURING OSMOTIC STRESS
Enterotoxigenic E. coli secrete heat stable enterotoxins, such as STa, that cause secretory diarrhea, an important cause of mortality in developing countries. Guanylin (Gn) and uroguanylin (Ugn) are endogenous peptide ligands that have significant homology to STa. Gn and Ugn are produced as prohormones in intestinal epithelia and are secreted and cleaved in the gut lumen. Like STa, these hormones activate guanylate cyclase C (GC-C) and cause intracellular levels of cGMP to increase. This eventually leads to phosphorylation of the cystic fibrosis transmembrane conductance regulator (CFTR) and subsequent ion flow from the cell. This dissertation seeks to test the hypothesis that Gn and Ugn play a substantial role in fluid secretion from intestinal epithelia and are important in controlling the hydration state of the intestinal contents. We used three model systems to address this hypothesis. First, a whole mouse model suggested that Gn and Ugn are not decreased during osmotic diarrhea as expected but are actually increased at RNA and protein levels. Second, an in vitro cell culture system confirmed that these genes are increased during hypertonic stress and further showed that compatible osmolytes can block induction of guanylin mRNA. In addition, this model implicated protein kinase C in the control of Gn and Ugn expression. Third, targeted inactivation of the mouse guanylin gene was accomplished. Guanylin null mice had normal weight gain and did not develop intestinal obstruction. This model suggested that these mice do not have a seriously deranged intestinal phenotype that is the result of diminished fluid secretion into the intestine. However, Gn null mice may have a reproductive or developmental phenotype and show subtle changes in intestinal crypt cell morphology. These three experimental systems indicate that the hypothesis concerning the role of guanylin and uroguanylin in gastrointestinal tract hydration needs to be revisited and modified. Increases in these ligands during hypertonic cellular water loss and lack of a severe phenotype in the Gn-/- mouse suggest that guanylin and uroguanylin have other functions in the intestine. If guanylin does play a role in intestinal fluid secretion, compensatory homeostatic mechanisms are able to correct for its loss.
School:University of Cincinnati
School Location:USA - Ohio
Source Type:Master's Thesis
Keywords:uroguanylin guanylate cyclase cre loxp mouse model
Date of Publication:01/01/2001