WNK1-OSR1 kinase-mediated phospho-activation of Na+-K+-2Cl- cotransporter facilitates glioma migration

Molecular Cancer

Research

Kelli Pointer¹  John S Kuo²  Kristopher T Kahle³  Paul A Clark⁴  Gulnaz Begum⁵  Wen Zhu⁵  Dandan Sun⁶  Shih-Hua Lin⁷  Sung-Sen Yang⁷ 

[1]Cellular and Molecular Biology Program, University of Wisconsin School of Medicine and Public Health, Madison, WI, USA

[2]Department of Neurological Surgery, University of Wisconsin School of Medicine and Public Health, Madison, WI, USA

[3]Cellular and Molecular Biology Program, University of Wisconsin School of Medicine and Public Health, Madison, WI, USA

[4]Department of Neurological Surgery, University of Wisconsin School of Medicine and Public Health, Madison, WI, USA

[5]Carbone Cancer Center, University of Wisconsin, Madison, WI, USA

[6]Department of Neurological Surgery, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA

[7]Department of Neurological Surgery, University of Wisconsin School of Medicine and Public Health, Madison, WI, USA

[8]Department of Neurology, University of Pittsburgh, Pittsburgh, PA, USA

[9]Department of Neurology, University of Pittsburgh, Pittsburgh, PA, USA

[10]Veterans Affairs Pittsburgh Health Care System, Geriatric Research, Educational and Clinical Center, Pittsburgh, PA, USA

[11]Department of Neurology, University of Pittsburgh, S-598 South Biomedical Science Tower (BST), 3500 Terrace Street, 15213, Pittsburgh, PA, USA

[12]Division of Nephrology, Department of Medicine, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan

关键词: Bumetanide;    Cell volume;    Ezrin;    Ion cotransporter;    Temozolomide;   

DOI  :  10.1186/1476-4598-13-31

 received in 2013-09-11, accepted in 2014-02-13,  发布年份 2014

来源: Springer

【 摘 要 】

BackgroundThe bumetanide (BMT)-sensitive Na+-K+-2Cl- cotransporter isoform 1 (NKCC1) maintains cell volume homeostasis by increasing intracellular K+ and Cl- content via regulatory volume increase (RVI). Expression levels of NKCC1 positively correlate with the histological grade and severity of gliomas, the most common primary adult brain tumors, and up-regulated NKCC1 activity facilitates glioma cell migration and apoptotic resistance to the chemotherapeutic drug temozolomide (TMZ). However, the cellular mechanisms underlying NKCC1 functional up-regulation in glioma and in response to TMZ administration remain unknown.MethodsExpression of NKCC1 and its upstream kinases With-No-K (Lysine) kinase 1 (WNK1) and oxidative stress-responsive kinase-1 (OSR1) in different human glioma cell lines and glioma specimens were detected by western blotting and immunostaining. Live cell imaging and microchemotaxis assay were applied to record glioma cell movements under different treatment conditions. Fluorescence indicators were utilized to measure cell volume, intracellular K+ and Cl- content to reflect the activity of NKCC1 on ion transportation. Small interfering RNA (siRNA)-mediated knockdown of WNK1 or OSR1 was used to explore their roles in regulation of NKCC1 activity in glioma cells. Results of different treatment groups were compared by one-way ANOVA using the Bonferroni post-hoc test in the case of multiple comparisons.ResultsWe show that compared to human neural stem cells and astrocytes, human glioma cells exhibit robust increases in the activation and phosphorylation of NKCC1 and its two upstream regulatory kinases, WNK1 and OSR1. siRNA-mediated knockdown of WNK1 or OSR1 reduces intracellular K+ and Cl- content and RVI in glioma cells by abolishing NKCC1 regulatory phospho-activation. Unexpectedly, TMZ activates the WNK1/OSR1/NKCC1 signaling pathway and enhances glioma migration. Pharmacological inhibition of NKCC1 with its potent inhibitor BMT or siRNA knockdown of WNK1 or OSR1 significantly decreases glioma cell migration after TMZ treatment.ConclusionTogether, our data show a novel role for the WNK1/OSR1/NKCC1 pathway in basal and TMZ-induced glioma migration, and suggest that glioma treatment with TMZ might be improved by drugs that inhibit elements of the WNK1/OSR1/NKCC1 signaling pathway.

【 授权许可】

Unknown   
© Zhu et al.; licensee BioMed Central Ltd. 2014. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. 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.