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. 2017 Jun 23;7:4149. doi: 10.1038/s41598-017-03510-5

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© The Author(s) 2017

Open Access This 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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.

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Mammalian two-hybrid demonstrates direct SGK1-mediated NCC and ENaC interaction. Mammalian two-hybrid assay performed on transiently transfected COS-7 cells with: empty vectors (pBIND, pACT), positive (Myo-pAct + Id-pBIND) and/or NCC-pBIND, αENaC (4A) or γ-pAct (4B), SGK1 or WNK4. Empty vector pCDNA was used to normalize for total amounts of DNA transfected. COS-7 cells lysed and values of Firefly and Renilla luciferase obtained. Firefly luciferase data were normalized to Renilla and all data expressed as fold change of negative control group (empty vector pBIND + pAct). Data represented as mean ± SEM, n = 7–12, at least 3 separate experiments: (a) ANOVA, *p < 0.05, **p < 0.01, ***p < 0.0001, (b) Kruskal-Wallis, **p < 0.01, ****p < 0.0001.