Expression and function of the guanylin ligand family

描述（由申请人提供）：观音肽和尿观音肽是主要在哺乳动物肠道中表达的肽，由于它们与细菌热稳定肠毒素（ST）的同源性以及它们与ST受体——观音酰环化酶C （GC-C）结合的能力而被鉴定出来。ST是世界性的分泌性腹泻的病因，而观音和尿观音被认为是调节肠道分泌而不引起腹泻。因此，更好地了解这些肽的作用机制和受体将有可能阐明产毒性腹泻的病理生理学。为了进一步探讨观音林和尿观音林的生理作用，我们建立了观音林和尿观音林基因靶向小鼠。我们在初步实验中发现尿观音在盐体内平衡中具有重要的调节作用。因此，本应用的首要假设是，尿观音和观音是肠道激素，在肠道中形成局部旁分泌调节系统和作为肠肾轴一部分调节尿钠的激素系统。具体目标如下：1)明确观音林和尿观音林作为调节基因靶向小鼠盐和水代谢的内分泌激素的作用。我们将验证以下假设：1)尿观音会导致尿钠和尿钾排泄受损，而不是观音或GC-C缺陷小鼠。2)尿观音林缺乏小鼠表现为盐敏感性高血压。3)尿观音的这些作用是通过一种新的非gc - c受体介导的。II)确定观音和尿观音表达的调控机制。我们将验证以下假设：1)观音碱和尿观音碱的基础表达水平受蛋白激酶C （PKC）亚型的调节。2)高渗透性通过转录调控、依赖pkc的机制上调观音和尿观音。3. 干扰素- γ (ifn - γ)通过转录调控机制下调观音和尿观音。III)明确尿观音在肠和肾中丢失导致的细胞分泌缺陷。我们将验证以下假设：1)尿观音缺失会导致肠道氯离子分泌受损。2)在肾脏中，尿观音的缺失会导致近端小管细胞的水和溶质运输失调，导致细胞内液泡形成。3)尿观音缺失会导致新型尿观音受体基因代偿性上调，以及小肠近端肠细胞和肾近端小管细胞中转运体和水通道表达的代偿性反调控。这些研究将进一步描述这些肽和ST在肠道中的作用和作用，以及它们在肠-肾轴中作为肠道激素的作用。此外，这些研究将为涉及这些肽作为液体过载状态治疗剂的转化研究奠定基础。

DESCRIPTION (provided by applicant): Guanylin and uroguanylin are peptides that are expressed primarily in the mammalian intestine and were identified because of their homology to the bacterial heat stable enterotoxin (ST) and their ability to bind to the ST receptor, guanylyl cyclase C (GC-C). ST is a worldwide cause of secretory diarrhea whereas guanylin and uroguanylin are thought to modulate intestinal secretion without causing diarrhea. Therefore, a better understanding of the mechanisms and receptors through which these peptides act will likely shed light on the pathophysiology of toxigenic diarrhea. To further explore the physiologic role of guanylin and uroguanylin we created guanylin and uroguanylin gene-targeted mice. We found in preliminary experiments that uroguanylin has an important regulatory role in salt homeostasis. Therefore, the overarching hypothesis of this application is that uroguanylin and guanylin are enteric hormones forming both a local paracrine regulatory system in the intestine and a hormonal system as part of an enteric-renal axis to regulate natriuresis. We will address the following specific aims. I) Define the role of guanylin and uroguanylin as endocrine hormones regulating salt and water metabolism in gene targeted mice. We will test the hypotheses that: 1) Uroguanylin but not guanylin or GC-C deficient mice, will demonstrate impaired urinary sodium and potassium excretion. 2) Uroguanylin deficient mice will demonstrate salt sensitive hypertension. 3) These actions of uroguanylin are mediated via a novel, non-GC-C receptor. II) Determine the mechanisms regulating guanylin and uroguanylin expression. We will test the hypotheses that: 1) Basal levels of guanylin and uroguanylin expression are regulated by protein kinase C (PKC) isoforms. 2) Hypertonicity upregulates guanylin and uroguanylin by a transcriptionally-regulated, PKC-dependent mechanism. 3. Interferon-gamma (IFN-gamma) downregulates guanylin and uroguanylin via a transcriptionally-regulated mechanism. III) Define the cellular secretory defects resulting from uroguanylin loss in intestine and kidney. We will test the hypotheses that: 1) Loss of uroguanylin will result in impaired intestinal secretion of chloride. 2) In the kidney, loss of uroguanylin will result in dysregulated water and solute transport in proximal tubular cells leading to intracellular vacuole formation. 3) Loss of uroguanylin will result in compensatory upregulation of novel uroguanylin receptor gene(s) and compensatory counter-regulation of transporters and water channels expressed in enterocytes in the proximal small intestine and proximal tubule cells in the kidney. These studies will further delineate the role and action of these peptides and ST in the intestine as well as their roles as intestinal hormones in an enteric-renal axis. Furthermore, these studies will lay the groundwork for translational research involving these peptides as therapeutic agents for fluid overload states.