Pendrin, a novel transcriptional target of the uroguanylin system.

Guanylin (GN) and uroguanylin (UGN) are low-molecular-weight peptide hormones produced mainly in the intestinal mucosa in response to oral salt load. GN and UGN (guanylin peptides) induce secretion of electrolytes and water in both intestine and kidney. Thought to act as "intestinal natriuretic factors", GN and UGN modulate renal salt secretion by both endocrine mechanisms (linking the digestive system and kidney) and paracrine/autocrine (intrarenal) mechanisms. The cellular function of GN and UGN in intestine and proximal tubule is mediated by guanylyl cyclase C (GC-C)-, cGMP-, and G protein-dependent pathways, whereas, in principal cells of the cortical collecting duct (CCD), these peptide hormones act via GC-C-independent signaling through phospholipase A2 (PLA2). The Cl(-)/HCO(-)3 exchanger pendrin (SLC26A4), encoded by the PDS gene, is expressed in non-α intercalated cells of the CCD. Pendrin is essential for CCD bicarbonate secretion and is also involved in NaCl balance and blood pressure regulation. Our recent studies have provided evidence that pendrin-mediated anion exchange in the CCD is regulated at the transcriptional level by UGN. UGN exerts an inhibitory effect on the pendrin gene promoter likely via heat shock factor 1 (HSF1) action at a defined heat shock element (HSE) site. Recent studies have unraveled novel roles for guanylin peptides in several organ systems including involvement in appetite regulation, olfactory function, cell proliferation and differentiation, inflammation, and reproductive function. Both the guanylin system and pendrin have also been implicated in airway function. Future molecular research into the receptors and signal transduction pathways involved in the action of guanylin peptides and the pendrin anion exchanger in the kidney and other organs, and into the links between them, may facilitate discovery of new therapies for hypertension, heart failure, hepatic failure and other fluid retention syndromes, as well as for diverse diseases such as obesity, asthma, and cancer.

The pendrin anion exchanger gene is transcriptionally regulated by uroguanylin: a novel enterorenal link.

The pendrin/SLC26A4 Cl(-)/HCO(3)(-) exchanger, encoded by the PDS gene, is expressed in cortical collecting duct (CCD) non-A intercalated cells. Pendrin is essential for CCD bicarbonate secretion and is also involved in NaCl balance and blood pressure regulation. The intestinal peptide uroguanylin (UGN) is produced in response to oral salt load and can function as an "intestinal natriuretic hormone." We aimed to investigate whether UGN modulates pendrin activity and to explore the molecular mechanisms responsible for this modulation. Injection of UGN into mice resulted in decreased pendrin mRNA and protein expression in the kidney. UGN decreased endogenous pendrin mRNA levels in HEK293 cells. A 4.2-kb human PDS (hPDS) promoter sequence and consecutive 5' deletion products were cloned into luciferase reporter vectors and transiently transfected into HEK293 cells. Exposure of transfected cells to UGN decreased hPDS promoter activity. This UGN-induced effect on the hPDS promoter occurred within a 52-bp region encompassing a single heat shock element (HSE). The effect of UGN on the promoter was abolished when the HSE located between nt -1119 and -1115 was absent or was mutated. Furthermore, treatment of HEK293 cells with heat shock factor 1 (HSF1) small interfering RNA (siRNA) reversed the UGN-induced decrease in endogenous PDS mRNA level. In conclusion, pendrin-mediated Cl(-)/HCO(3)(-) exchange in the renal tubule may be regulated transcriptionally by the peptide hormone UGN. UGN exerts its inhibitory activity on the hPDS promoter likely via HSF1 action at a defined HSE site. These data define a novel signaling pathway involved in the enterorenal axis controlling electrolyte and water homeostasis.

Transcriptional regulation of the pendrin gene.

Pendrin (SLC26A4), a Cl(-)/anion exchanger encoded by the gene PDS, is highly expressed in the kidney, thyroid and inner ear epithelia and is essential for bicarbonate secretion/chloride reabsorption, iodide accumulation and endolymph ion balance, respectively. The molecular mechanisms controlling pendrin activity in renal, thyroid and inner ear epithelia have been the subject of recent studies. The effects of ambient pH, the hormone aldosterone and the peptide uroguanylin (UGN; the "intestinal natriuretic hormone"), known modulators of electrolyte balance, on transcription of the pendrin gene, have been investigated. Luciferase reporter plasmids containing different length fragments of the human PDS (hPDS) promoter were transfected into renal HEK293, thyroid LA2, and inner ear VOT36 epithelial cells. Acidic pH decreased and alkaline pH increased hPDS promoter activity in transfected HEK293 and VOT36, but not in LA2 cells. Aldosterone reduced hPDS promoter activity in HEK293 but had no effect in LA2 and VOT36 cells. These pH and aldosterone-induced effects on the hPDS promoter occurred within 96-bp and 89-bp regions, respectively, which likely contain distinct response elements to these modulators. Injection of UGN into mice resulted in decreased pendrin mRNA and protein expression in the kidney. Exposure of transfected HEK293 to UGN decreased hPDS promoter activity. The findings provided evidence for the presence of a UGN response element within the 96-bp region overlapping with the pH response element on the hPDS promoter. Pendrin is also expressed in airway epithelium. The cytokins interleukin 4 (IL-4) and interleukin-13 (IL-13), known regulators of airway surface function, have been shown to increase hPDS promoter activity by a STAT6-dependent mechanism. In conclusion, systemic pH, the hormone aldosterone, and the peptide UGN influence renal tubular pendrin gene expression and, perhaps, pendrin-mediated Cl(-)/HCO(3)(-) exchange at the transcriptional level. Pendrin-driven anion transport in the endolymph and at the airway surface may be regulated transcriptionally by systemic pH and IL-3/IL-4, respectively. The distinct response elements and the corresponding transcription factors mediating the effect of these modulators on the PDS promoter remain to be identified and characterized.

Guanylin and its lysine-containing analogue in the isolated perfused rat kidney: interaction with chymotrypsin inhibitor.

Guanylin and uroguanylin are two novel peptides that activate membrane-bound guanylate cyclases found in the kidney and intestine, influencing fluid and electrolyte homeostasis by cyclic GMP. Their natriuretic and kaliuretic activities are well documented. Since guanylin is inactivated by chymotrypsin in vitro, experiments were designed to evaluate the role of chymotrypsin-like proteases in renal metabolism of guanylin. Using the isolated perfused rat kidney, guanylin and a recombinant derivative containing a lysine residue in the N-terminus of the native peptide was tested. There were three experimental groups. In the first group, lys-guanylin (0.1-2.5 microg/ml) was placed into perfusate reservoir. In the second group, chymostatin (6 microg/ml), a chymotrypsin inhibitor, was placed into solution. In the third group, after 30 min. of perfusion with chymostatin (6 microg/ml), guanylin (0.3 microg/ml) was placed into solution. A maximal decrease in fractional Na+ reabsorption (%TNa+) was achieved at 1.0 microg/ml of lys-guanylin (from 73.25+/-2.29 to 54.97+/-0.10, P<0.05). Lys-guanylin (1.0 microg/ml) also decreased fractional K+ reabsorption (%TK+) from 59.26+/-3.93 to 30.75+/-0.78 (P<0.05). Chymostatin had no detectable effects in electrolyte reabsorption in this assay. When introduced after chymostatin, guanylin lowered %TNa+ (from 81.2+/-1.86 to 72.6+/-2.45, P<0.05) and %TK+ (from 69.4+/-4.12 to 65.8+/-2.81, P<0.05). At this subthreshold concentration, guanylin alone lacks effects in %TNa+ or %TK+. Furthermore, the ability of both peptides to promote increases in intestinal fluid secretion was evaluated in the in vivo suckling mouse model. When administered per os, guanylin failed to stimulate intestinal secretion. When chymostatin was present in the test solution, guanylin induced intestinal secretion in this assay. In marked contrast, lys-guanylin alone induced diarrhoea in the suckling mouse. The present paper concludes that guanylin undergoes metabolism in target tissues such as the intestine and kidney and its lysine-containing analogue retains full biological activity.

Guanylin and uroguanylin induce natriuresis in mice lacking guanylyl cyclase-C receptor.

BACKGROUND: Guanylin (GN) and uroguanylin (UGN) are intestinally derived peptide hormones that are similar in structure and activity to the diarrhea-causing Escherichia coli heat-stable enterotoxins (STa). These secretagogues have been shown to affect fluid, Na+, K+, and Cl- transport in both the intestine and kidney, presumably by intracellular cyclic guanosine monophosphate (cGMP)-dependent signal transduction. However, the in vivo consequences of GN, UGN, and STa on renal function and their mechanism of action have yet to be rigorously tested. METHODS: We hypothesized that intravenous administration of GN, UGN, or STa would cause an increase in natriuresis in wild-type mice via cGMP and guanylyl cyclase-C (GC-C, Gucy2c), the only known receptor for these peptide-hormones, and that the peptide-induced natriuresis would be blunted in genetically altered mice devoid of GC-C receptors (GC-C(-/-) null). RESULTS: In wild-type mice using a modified renal clearance model, GN, UGN, and STa elicited significant natriuresis, kaliuresis, and diuresis as well as increased urinary cGMP levels in a time- and dose-dependent fashion. Absolute and fractional urinary sodium excretion levels were greatest approximately 40 minutes following a bolus infusion with pharmacologic doses of these peptides. Unexpectedly, GC-C(-/-) null mice also responded to the GN peptides similarly to that observed in wild-type mice. Glomerular filtration rate (GFR), blood pressure, and plasma cGMP in the mice (wild-type or GC-C(-/-) null) did not significantly vary between the vehicle- and peptide-treatment groups. The effects of UGN may also influence long-term renal function due to down-regulation of the Na+/K+ ATPase gamma-subunit and the Cl- channel ClC-K2 by 60% and 75%, respectively, as assessed by differential display polymerase chain reaction (PCR) (DD-PCR) and Northern blot analysis of kidney mRNA from mice treated with UGN. CONCLUSION: GN, UGN, and STa act on the mouse kidney, in part, through a cGMP-dependent, GC-C-independent mechanism, causing significant natriuresis by renal tubular processes. UGN may have further long-term effects on the kidney by altering the expression of such transport-associated proteins as Na+/K+ ATPase and ClC-K2.