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Renal arginine vasopressin receptor 2 (AVPR2) plays a crucial role in osmoregulation. Engagement of ligand with AVPR2 results in aquaporin 2 movement to the apical membrane and water reabsorption from the urinary filtrate. Despite this essential role, little is known about transcriptional regulation of Avpr2. Here, we identify novel roles for PAX2, a transcription factor crucial for kidney development, and its adaptor protein, Pax transcription interacting protein (PTIP), for epigenetic regulation of Avpr2 and thus body water balance. Chromatin immunoprecipitation (ChIP) from murine inner medulla cells (IMCD-3) identified the minimal DNA-binding region of PAX2 on the Avpr2 promoter. Regulation of Avpr2 by PAX2 was confirmed using a heterologous DNA expression system. PAX2 recruits the adaptor protein PTIP and its associated histone methyltransferase (HMT) complex to Avpr2 promoter, imposing epigenetic marks on this region and throughout the coding sequence that modulate Avpr2 gene transcription. Reduction of PAX2 or PTIP protein levels by siRNA prevented histone lysine methylation and expression of Avpr2. ChIP using mouse or human kidneys determined that PAX2 is highly enriched in the AVPR2 promoter alongside PTIP and HMT proteins, leading to high levels of histone H3 lysine trimethylation within the promoter and throughout the gene. In conclusion, PAX2 provides locus specificity for PTIP, allowing the HMT complex to impart epigenetic changes at the Avpr2 locus and regulate Avpr2 transcription. These finding have major implications for understanding regulation of body water balance. NEW & NOTEWORTHY The transcription factor PAX2 plays an indispensable role in kidney development. In the adult kidney, we identified the first described protein this protein regulates. PAX2 and its interacting partner Pax transcription interacting protein recruit a histone methyltransferase complex to the promoter and epigentically regulate the expression of arginine vasopressin receptor 2, a protein that plays a crucial role in osmoregulation in the distal tubule. [ABSTRACT FROM AUTHOR]

This article challenges both contemporary and classic urban theory by analyzing the historical case of coastal Ecuador. Working from primary and secondary sources, I track the urbanization of coastal Ecuador during the long nineteenth century, when cacao exports determined not only the economic wellbeing of the city of Guayaquil, but of the entire tropical lowland region. I argue that this extended urban geography was both experienced and practiced as an unbounded economic and cultural region. As the value of cacao exports skyrocketed, capitalists in the city invested in infrastructural projects and financial instruments, divorcing money‐making from cacao production. After the Gran Incendio (great fire) of 1896, the city was rebuilt according to the ideals of modern liberal planning that further separated the city from the country symbolically, despite their continued material interconnection. This work suggests that long histories of capitalist urbanization provide material and theoretical support for critiquing bounded urban theory both past and present, by moving beyond the city and highlighting the processes undergirding spatial production under capitalist social relations. Likewise, this historical case study argues that city‐centrism, rather than being constituted epistemologically, was tied to liberal notions of the urban based on nineteenth‐century ideologies of modernization and progress. [ABSTRACT FROM AUTHOR]

Background: Furosemide inhibits the sodium potassium chloride cotransporter (NKCC2) in the thick ascending limb of the loop of Henle and increases urinary water and sodium excretion. This study investigates the effect of furosemide on body composition estimated with multifrequency bioimpedance spectroscopy (BIS) technique and urinary proteins from NKCC2. Methods: This study is a randomized, placebo‐controlled, crossover study where healthy subjects received either placebo or 40 mg furosemide on two separate occasions, where body composition with BIS, renal function, proteins from tubular proteins that mediate sodium and water transport, and plasma concentrations of vasoactive hormones were measured before and after intervention. Results: We observed an expected increased diuresis with a subsequent reduction in bodyweight of (−1.51 ± 0.36 kg, p <.001) and extracellular water (ECW; −1.14 ± 0.23 L, p <.001) after furosemide. We found a positive correlation between the decrease in ECW and a decrease in bodyweight and a negative correlation between the decrease in ECW and the increase in urinary output. Intracellular water (ICW) increased (0.47 ± 0.28 L, p <.001). Urinary excretion of NKCC2 increased after furosemide and the increase in NKCC2 correlated with an increase in urine output and a decrease in ECW. Conclusion: We found BIS can detect acute changes in body water content but the method may be limited to estimation of ECW. BIS demonstrated that furosemide increases ICW which might be explained by an extracellular sodium loss. Finally, urinary proteins from NKCC2 increases after furosemide with a good correlation with diuresis end the decrease in ECW. [ABSTRACT FROM AUTHOR]

The sodium-hydrogen exchanger isoform 3 (NHE3, SLC9A3) is abundantly expressed in the gastrointestinal tract and is proposed to play essential roles in Na+ and fluid absorption as well as acid-base homeostasis. Mutations in the SLC9A3 gene can cause congenital sodium diarrhea (CSD). However, understanding the precise role of intestinal NHE3 has been severely hampered due to the lack of a suitable animal model. To navigate this problem and better understand the role of intestinal NHE3, we generated a tamoxifen-inducible intestinal epithelial cell-specific NHE3 knockout mouse model (NHE3IEC-KO). Before tamoxifen administration, the phenotype and blood parameters of NHE3IEC-KO were unremarkable compared with control mice. After tamoxifen administration, NHE3IEC-KO mice have undetectable levels of NHE3 in the intestine. NHE3IEC-KO mice develop watery, alkaline diarrhea in combination with a swollen small intestine, cecum and colon. The persistent diarrhea results in higher fluid intake. After 3 weeks, NHE3IEC-KO mice show a ~25% mortality rate. The contribution of intestinal NHE3 to acid-base and Na+ homeostasis under normal conditions becomes evident in NHE3IEC-KO mice that have metabolic acidosis, lower blood bicarbonate levels, hyponatremia and hyperkalemia associated with drastically elevated plasma aldosterone levels. These results demonstrate that intestinal NHE3 has a significant contribution to acid-base, Na+ and volume homeostasis, and lack of intestinal NHE3 has consequences on intestinal structural integrity. This mouse model mimics and explains the phenotype of individuals with CSD carrying SLC9A3 mutations. [ABSTRACT FROM AUTHOR]

After the release of bilateral ureteral obstruction (BUO), postobstructive diuresis from an impaired urine concentration mechanism is associated with reduced aquaporin 2 (AQP2) abundance in the inner medullary collecting duct (IMCD). However, the underlying molecular mechanism of this AQP2 reduction is incompletely understood. To elucidate the mechanisms responsible for this phenomenon, we studied molecular changes in IMCDs isolated from rats with 4-h BUO or sham operation at the early onset of AQP2 downregulation using mass spectrometry-based proteomic analysis. Two-hundred fifteen proteins had significant changes in abundances, with 65% of them downregulated in the IMCD of 4-h BUO rats compared with sham rats. Bioinformatic analysis revealed that significantly changed proteins were associated with functional Gene Ontology terms, including "cell-cell adhesion," "cell-cell adherens junction," "mitochondrial inner membrane," "endoplasmic reticulum chaperone complex," and the KEGG pathway of glycolysis/gluconeogenesis. Targeted liquid chromatography-tandem mass spectrometry or immunoblot analysis confirmed the changes in 19 proteins representative of each predominant cluster, including AQP2. Electron microscopy demonstrated disrupted tight junctions, disorganized adherens junctions, swollen mitochondria, enlargement of the endoplasmic reticulum lumen, and numerous autophagosomes/lysosomes in the IMCD of rats with 4-h BUO. AQP2 and seven proteins chosen as representative of the significantly altered clusters had a significant increase in immunofluorescence-based colocalization with autophagosomes/lysosomes. Immunogold electron microscopy confirmed colocalization of AQP2 with the autophagosome marker microtubule-associated protein 1A/1B-light chain 3 and the lysosomal marker cathepsin D in IMCD cells of rats with 4-h BUO. We conclude that enhanced autophagic degradation of AQP2 and other critical proteins, as well as endoplasmic reticulum stress in the IMCD, are initiated shortly after BUO. [ABSTRACT FROM AUTHOR]

The metabolic sensor AMP-activated protein kinase (AMPK) inhibits the epithelial Na+ channel (ENaC), a key regulator of salt reabsorption by the kidney and thus total body volume and blood pressure. Recent studies have suggested that AMPK promotes the association of p21-activated kinase-interacting exchange factor-ß1 ß1Pix, 14-3-3 proteins, and the ubiquitin ligase neural precursor cell expressed developmentally downregulated protein (Nedd)4-2 into a complex that inhibits ENaC by enhancing Nedd4-2 binding to ENaC and ENaC degradation. Functional ß1Pix is required for ENaC inhibition by AMPK and promotes Nedd4-2 phosphorylation and stability in mouse kidney cortical collecting duct cells. Here, we report that AMPK directly phosphorylates ß1Pix in vitro. Among several AMPK phosphorylation sites on ß1Pix detected by mass spectrometry, Ser71 was validated as functionally significant. Compared with wild-type ß1Pix, overexpression of a phosphorylation-deficient ß1Pix-S71A mutant attenuated ENaC inhibition and the AMPK-activated interaction of both ß1Pix and Nedd4-2 to 14-3-3 proteins in cortical collecting duct cells. Similarly, overexpression of a ß1Pix-A602-611 deletion tract mutant unable to bind 14-3-3 proteins decreased the interaction between Nedd4-2 and 14-3-3 proteins, suggesting that 14-3-3 binding to ß1Pix is critical for the formation of a ß1Pix/Nedd4-2/14-3-3 complex. With expression of a general peptide inhibitor of 14-3-3-target protein interactions (R18), binding of both ß1Pix and Nedd4-2 to 14-3-3 proteins was reduced, and AMPK-dependent ENaC inhibition was also attenuated. Altogether, our results demonstrate the importance of AMPK-mediated phosphorylation of ß1Pix at Ser71, which promotes 14-3-3 interactions with ß1Pix and Nedd4-2 to form a tripartite ENaC inhibitory complex, in the mechanism of ENaC regulation by AMPK. [ABSTRACT FROM AUTHOR]