ArticlePloS one2011
Phosphorylation and transport in the Na-K-2Cl cotransporters, NKCC1 and NKCC2A, compared in HEK-293 cells.
Article in PloS one, 2011. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 24 papers.
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Who cites it
24 citing papers in PubMed, 37 citations in OpenAlex.
- Reactive Oxygen Species (ROS) in Metabolic Disease-Don't Shoot the Metabolic Messenger.International journal of molecular sciences · 2025Review
- Update on NKCC2 regulation in the thick ascending limb (TAL) by membrane trafficking, phosphorylation, and protein-protein interactions.Frontiers in physiology · 2024Review
- The role ofJournal of pharmaceutical analysis · 2023Review
- Tubuloid culture enables long-term expansion of functional human kidney tubule epithelium from iPSC-derived organoids.Proceedings of the National Academy of Sciences of the United States of America · 2023Article
- The Post-Translational Modification Networking in WNK-Centric Hypertension Regulation and Electrolyte Homeostasis.Biomedicines · 2022Review
- Targeting the WNK-SPAK/OSR1 Pathway and Cation-Chloride Cotransporters for the Therapy of Stroke.International journal of molecular sciences · 2021Review
- Glial Chloride Homeostasis Under Transient Ischemic Stress.Frontiers in cellular neuroscience · 2021Article
- The role of WNK in modulation of KCl cotransport activity in red cells from normal individuals and patients with sickle cell anaemia.Pflugers Archiv : European journal of physiology · 2019Article
- A role for KCC3 in maintaining cell volume of peripheral nerve fibers.Neurochemistry international · 2019Review
- Cytokine-Regulation of NaFrontiers in immunology · 2017Review
- SPAK and OSR1 play essential roles in potassium homeostasis through actions on the distal convoluted tubule.The Journal of physiology · 2016Article
- Increased Slc12a1 expression in β-cells and improved glucose disposal in Slc12a2 heterozygous mice.The Journal of endocrinology · 2015Article
- A unifying mechanism for WNK kinase regulation of sodium-chloride cotransporter.Pflugers Archiv : European journal of physiology · 2015Review
- STE20/SPS1-related proline/alanine-rich kinase (SPAK) is critical for sodium reabsorption in isolated, perfused thick ascending limb.American journal of physiology. Renal physiology · 2015Article
- Ammonia, like K(+), stimulates the Na(+), K(+), 2 Cl(-) cotransporter NKCC1 and the Na(+),K(+)-ATPase and interacts with endogenous ouabain in astrocytes.Neurochemical research · 2015Review
- Impact of Hybrid and Complex N-Glycans on Cell Surface Targeting of the Endogenous Chloride Cotransporter Slc12a2.International journal of cell biology · 2015Article
- Identification of the WNK-SPAK/OSR1 signaling pathway in rodent and human lenses.Investigative ophthalmology & visual science · 2014Article
- Regulation of NKCC2 activity by inhibitory SPAK isoforms: KS-SPAK is a more potent inhibitor than SPAK2.American journal of physiology. Renal physiology · 2013Article
- Interactions with WNK (with no lysine) family members regulate oxidative stress response 1 and ion co-transporter activity.The Journal of biological chemistry · 2012Article
- Activation of thiazide-sensitive co-transport by angiotensin II in the cyp1a1-Ren2 hypertensive rat.PloS one · 2012Article
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Authors and funding
2 authors at 1 institution in 1 country.
Funding
Abstract
Na-K-2Cl cotransporters help determine cell composition and volume. NKCC1 is widely distributed whilst NKCC2 is only found in the kidney where it plays a vital role reabsorbing 20% of filtered NaCl. NKCC2 regulation is poorly understood because of its restricted distribution and difficulties with its expression in mammalian cell cultures. Here we compare phosphorylation of the N-termini of the cotransporters, measured with phospho-specific antibodies, with bumetanide-sensitive transport of K(+) ((86)Rb(+)) (activity) in HEK-293 cells stably expressing fNKCC1 or fNKCC2A which were cloned from ferret kidney. Activities of transfected transporters were distinguished from those of endogenous ones by working at 37 °C. fNKCC1 and fNKCC2A activities were highest after pre-incubation of cells in hypotonic low-[Cl(-)] media to reduce cell [Cl(-)] and volume during flux measurement. Phosphorylation of both transporters more than doubled. Pre-incubation with ouabain also strongly stimulated fNKCC1 and fNKCC2A and substantially increased phosphorylation, whereas pre-incubation in Na(+)-free media maximally stimulated fNKCC1 and doubled its phosphorylation, but inhibited fNKCC2A, with a small increase in its phosphorylation. Kinase inhibitors halved phosphorylation and activity of both transporters whereas inhibition of phosphatases with calyculin A strongly increased phosphorylation of both transporters but only slightly stimulated fNKCC1 and inhibited fNCCC2A. Thus kinase inhibition reduced phosphorylation and transport, and transport stimulation was only seen when phosphorylation increased, but transport did not always increase with phosphorylation. This suggests phosphorylation of the N-termini determines the transporters' potential capacity to move ions, but final activity also depends on other factors. Transport cannot be reliably inferred solely using phospho-specific antibodies on whole-cell lysates.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.