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Abstract: PO1413

Architecture of the Distal Nephron Mineralocorticoid Receptor-Dependent Transcriptome Defined

Session Information

Category: Fluid, Electrolyte, and Acid-Base Disorders

  • 901 Fluid, Electrolyte, and Acid-Base Disorders: Basic

Authors

  • Jung, Hyun Jun, Johns Hopkins University School of Medicine, Baltimore, Maryland, United States
  • Su, Xiao-Tong, Oregon Health and Science University, Portland, Oregon, United States
  • Al-Qusairi, Lama, Johns Hopkins University School of Medicine, Baltimore, Maryland, United States
  • Ellison, David H., Oregon Health and Science University, Portland, Oregon, United States
  • Welling, Paul A., Johns Hopkins University School of Medicine, Baltimore, Maryland, United States
Background

The mineralocorticoid receptor (MR, Nr3c2) is responsible for aldosterone-regulation of Na+ and K+ balance and blood pressure. Although a handful of aldosterone/MR-dependent genes have been identified, their regulation cannot fully explain how aldosterone activates electrogenic Na+- K+ exchange in the aldosterone sensitive distal nephron (ASDN). Here, we apply RNA-Seq and bioinformatic approaches in isolated tubule segments of MR KO vs. Control mice to define a more complete inventory of MR-dependent genes.

Methods

MRfl/fl/Pax8-rtTA/LC1 mice were used as a doxycycline (DOX)-inducible Nr3c2 gene KO model. After DOX treatment, four groups were prepared to distinguish between K+ and MR effects: 1) control mice on normal K+ diet (CT-NK) or 2) high K+ diet (CT-HK) and 3) MR knockout mice on normal K+ diet (KO-NK) or 4) low K+ diet (KO-LK). RNA-Seq analysis was carried out in the micro-dissected connecting tubule and cortical collecting duct tubule segments (5-6 mice per group and ~10 fresh ASDN tubules per mouse). Differential expression (DE) genes were identified (FDR < 0.05) and used for further bioinformatic analyses.

Results

927 and 2010 DE genes were identified from comparisons of MR KO-NK vs. CT-NK and MR KO-LK vs. CT-HK, respectively. Diet effects were not detected. Absence of transcripts on the third exon of Nr3c2 gene confirmed complete disruption of Nr3c2 gene in the MR KO. All known aldosterone-response genes, including Sgk1, Scnn1a, Ndrg2, Per3, Tsc22d3, Zbtb16, Mlph and Atp1a1 were significantly decreased in MR KO-LK compared to CT-HK. In addition, 5 DE genes (Sgk1, Scnn1a, Nedd4l, Fxyd4 and Atp1a1) were mapped on small known “Aldosterone-regulated sodium reabsorption” profile. However, genome-wide identification of GR and MR binding sites revealed that 526 of the significantly down-regulated genes in MR KO mice are potential MR-regulated genes. Pathway enrichment analysis of 2010 DE genes showed that DE genes were highly enriched in mitochondria-associated metabolic processes.

Conclusion

The inventory of MR-regulated genes in the ASDN is much larger than previously imagined. In addition to pathways that directly up-regulate epithelial sodium channel (ENaC) and the Na+-K+ ATPase, the data suggest that aldosterone-MR may directly influence metabolism to make energy-consuming transport highly efficient.

Funding

  • NIDDK Support