Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2021 Jul 7;40:226. doi: 10.1186/s13046-021-01982-y

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© The Author(s) 2021

Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

PMC Copyright notice

Fig. 1 — Hyperosmolality-induced upregulation of Ranbp3l is abrogated in NFAT5 deficient cells. A RNA-Seq analyses show the expression of Ranbp3l and other Ranbp family members in primary murine IMCD-cells cultivated at 300 or 600 msomol/kg displayed as heatmap in FPKM (Fragments per kilo base per million mapped reads), (n = 2). Blue to red represents lower to higher expression. B Pearson correlation analysis of genes affected by hyperosmolality (log₂ fold change > 1, <− 1) between the mpkCCD cell line and primary cultivated IMCD cells. (n = 2–4), blue = overlapping upregulated genes, red = overlapping down regulated genes. C WT (wildtype) mpKCCD cells were cultivated under isosmotic and hyperosmotic conditions for 7d and subjected to immunofluorescence staining with anti-NFAT5 antibody and DAPI for nucleus visualization. Scale bar: 100 μm (D) Western blot analysis of NFAT5 expression in scramble (Scr) and NFAT5 deficient single cell clones N1 and N3. The expression of GAPDH served as loading control. E The relative expression of Ranbp3l, Aqp2 and Slc6a12 in Scr vs. NFAT5-deficient clones was analyzed by RT-qPCR in cells cultivated under hyperosmotic conditions (n = 3). Values represent mean ± SEM (error bars). *, p < 0.05; **, p < 0.01, 1-way ANOVA. F Efficient in vitro deletion of NAFT5 in primary cultured IMCD cells using Nfat5^flx/fl-Ubc-Cre-ERT2^+/− mice. Representative immunofluorescence of control and 4-OH-TM treated cells for NFAT5 under hyperosmolar cell cultivation. Scale bar: 100 μm (G) FPKM values of all known kidney-specific NFAT5 targets, including Ranbp3l for control and 4-OH-TM treated cells at 300 mosmol/kg and 600 mosmol/kg (n = 2). (H) Induction of promotor activity under hyperosmotic conditions with the Ranbp3l-2 kb construct. (n = 4, each with duplicates). **, p < 0.01, Student’s t test (I) Luciferase assay with different Ranbp3l promoter fragments. Promoter activity is represented relative to the Ranbp3l-2 kb construct. (n = 4, with two technical replicates) n.s. > 0.05, *, p < 0.05, 1way ANOVA