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. 2020 Jan 7;11:78. doi: 10.1038/s41467-019-13851-6

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© The Author(s) 2020

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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Fig. 2 — a ZT-1a dose dependently inhibited KCC3 Thr991/Thr1048 phosphorylation in cells. HEK-293 cells were transfected with a DNA construct encoding wild-type N-terminally FLAG-tagged KCC3. Cells at 36 h post transfection were exposed for 30 min to either control isotonic or hypotonic low [Cl⁻] conditions, and then treated in the same conditions for an additional 30 min in the presence of inhibitors at the indicated concentrations. Cell lysates were subjected to SDS-PAGE and immunoblot with the indicated antibodies. b Immunoblot quantitation on ZT-1a, Closantel, STOCK1S-14279, and STOCK1S-50699. Band intensities were quantitated with ImageJ software. ***p < 0.001; **p < 0.01; *p < 0.05; ns, nonsignificant [by one-way ANOVA with post hoc testing (n = 6, error bars represent the mean ± SEM)].