Genetic deletion of the nuclear factor of activated T cells 5 in collecting duct principal cells causes nephrogenic diabetes insipidus

Abstract

Water homeostasis is tightly regulated by the kidneys via the process of urine concentration. During reduced water intake, the antidiuretic hormone arginine vasopressin (AVP) binds to the vasopressin receptor type II (V2R) in the kidney to enhance countercurrent multiplication and medullary osmolality, and increase water reabsorption via aquaporin-2 (AQP2) water channels. The importance of this AVP, V2R, and AQP2 axis is highlighted by low urine osmolality and polyuria in people with various water balance disorders, including nephrogenic diabetes insipidus (NDI). ELF5 and nuclear factor of activated T cells 5 (NFAT5) are two transcription factors proposed to regulate Aqp2 expression, but their role is poorly defined. Here we generated two novel mouse lines with principal cell (PC)-specific deletion of ELF5 or NFAT5 and phenotyped them in respect to renal water handling. ELF5-deficient mice (ELF5 PC-KO ) had a very mild phenotype, with no clear differences in AQP2 abundance, and mild differences in renal water handling and maximal urinary concentrating capacity. In contrast, NFAT5 (NFAT5 PC-KO ) mice had significantly higher water intake and their 24 h urine volume was almost 10fold greater than controls. After challenging with dDAVP or 8 h water restriction, NFAT5 PC-KO mice were unable to concentrate their urine, demonstrating that they suffer from NDI. The abundance of AQP2, other AQPs, and the urea transporter UT-A1 were greatly decreased in NFAT5 PC-KO mice. In conclusion, NFAT5 is a major regulator of not only Aqp2 gene transcription, but also other genes important for water homeostasis and its absence leads to the development of NDI.

Differences in chromatin accessibility between renal cortex and inner medulla correlate with spatial differences in gene expression and are modulated by NFAT5 function

“…ELF5 is a principal cell specific TF and might be involved in Aqp2 expression 55 . However, a collecting duct principal cell specific ELF5 deficient mice showed only minor differences in Aqp2 expression 32 .…”

“…Surprisingly the analysis of TF motifs did not show a significant enrichment of NFAT5 motifs in the IM for DAR regions. Loss of NFAT5 in the induced a diabetes insipidus like phenotype and this was associated with massive changes in gene expression 29, 32 . The changes in gene expression are also associated with differences in chromatin accessibility.…”

“…Several studies using cell lines and native tissue have shown, that hypertonicity and NFAT5 are master regulators of Aqp2 expression 29, 32, 52, 65 . Here we showed that the increased Aqp2 expression in the IM is associated with a gain of accessibility of Aqp2 at the promotor region.…”

“…To generate mice (strain C57BL/6N) that are deficient in NFAT5 in the principal cell (PC) of the collecting duct, floxed NFAT5 mice (NFAT5-fl/fl) were crossed with Aqp2-Cre mice expressing CRE recombinase under the control of the Aqp2 gene promoter 29, 32 . The insertion of the Cre recombinase disrupts one allele of the Aqp2 gene.…”

“…Our data show that chromatin accessibility and the effect of NFAT5 play an important role in influencing the gene expression profile. The NFAT5-KO mice exhibit a nephrogenic diabetes insipidus-like phenotype that is likely due to decreased expression of AQP2 32 . In primary cultured IMCD cells, the expression of Aqp2 is strongly induced under hypertonic cell culture conditions 28 .…”