Expression and function of the guanylin ligand family

描述（由申请人提供）：鸟苷素和尿鸟苷素是主要在哺乳动物肠道中表达的肽，由于其与细菌热稳定肠毒素（ST）的同源性及其与ST受体鸟苷酸环化酶C（GC-C）结合的能力而被鉴定。ST是分泌性腹泻的全球性原因，而鸟苷素和尿鸟苷素被认为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可能揭示致炎性腹泻的病理生理学。为了进一步探索鸟苷素和尿鸟苷素的生理作用，我们创建了鸟苷素和尿鸟苷素基因靶向小鼠。我们在初步实验中发现，尿鸟苷素在盐稳态中具有重要的调节作用。因此，本申请的首要假设是尿鸟苷素和鸟苷素是肠激素，其形成肠中的局部旁分泌调节系统和作为肠-肾轴的一部分的激素系统以调节尿钠排泄。我们将致力于实现以下具体目标。I）定义鸟苷素和尿鸟苷素作为调节基因靶向小鼠中盐和水代谢的内分泌激素的作用。我们将检验以下假设：1）尿鸟苷素而非鸟苷素或GC-C缺陷小鼠将显示尿钠和钾排泄受损。2)尿鸟苷素缺乏小鼠将表现出盐敏感性高血压。3)尿鸟苷素的这些作用是通过一种新的非GC-C受体介导的。II）确定调节鸟苷素和尿鸟苷素表达的机制。我们将检验以下假设：1）鸟苷素和尿鸟苷素表达的基础水平受蛋白激酶C（PKC）亚型的调节。2)高渗通过转录调节的PKC依赖性机制上调鸟苷素和尿鸟苷素。3.干扰素-γ（IFN-γ）通过转录调节机制下调鸟苷素和尿鸟苷素。III）定义肠和肾中由尿鸟苷素损失引起的细胞分泌缺陷。我们将检验以下假设：1）尿鸟苷素的缺失将导致肠氯化物分泌受损。2)在肾脏中，尿鸟苷素的损失将导致近端肾小管细胞中的水和溶质转运失调，导致细胞内空泡形成。3)尿鸟苷素的损失将导致新的尿鸟苷素受体基因的补偿性上调和近端小肠中的肠细胞和肾脏中的近端小管细胞中表达的转运蛋白和水通道的补偿性反调节。这些研究将进一步阐明这些肽和ST在肠道中的作用和作用，以及它们作为肠激素在肠-肾轴中的作用。此外，这些研究将为涉及这些肽作为液体超负荷状态的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.