Tyrosine phosphorylation modulates cell surface expression of chloride cotransporters NKCC2 and KCC3
Authors: Cl√°udia Almeida Loureiro
Patr√≠cia Barros
Paulo Matos
Peter Jordan
Journal: Archives of Biochemistry and Biophysics (2019)
Discipline: BIOLOGY & BIOCHEMISTRY
Keywords: Chloride cotransport; KCC3; Membrane traffic; NKCC2; Protein phosphorylation
Affiliations: Department of Human Genetics, National Health Institute 'Dr. Ricardo Jorge', Lisbon, Portugal; BioISI - Biosystems & Integrative Sciences Institute, Faculty of Sciences, University of Lisbon, Portugal., Department of Human Genetics, National Health Institute 'Dr. Ricardo Jorge', Lisbon, Portugal; BioISI - Biosystems & Integrative Sciences Institute, Faculty of Sciences, University of Lisbon, Portugal; Department of Chemistry and Biochemistry, Faculty of Sciences, University of Lisbon, Portugal., Department of Human Genetics, National Health Institute 'Dr. Ricardo Jorge', Lisbon, Portugal; BioISI - Biosystems & Integrative Sciences Institute, Faculty of Sciences, University of Lisbon, Portugal. Electronic address: .

Abstract:  Cellular chloride transport has a fundamental role in cell volume regulation and renal salt handling. Cellular chloride entry or exit are mediated at the plasma membrane by cotransporter proteins of the solute carrier 12 family. For example, NKCC2 resorbs chloride with sodium and potassium ions at the apical membrane of epithelial cells in the kidney, whereas KCC3 releases chloride with potassium ions at the basolateral membrane. Their ion transport activity is regulated by protein phosphorylation in response to signaling pathways. An additional regulatory mechanism concerns the amount of cotransporter molecules inserted into the plasma membrane. Here we describe that tyrosine phosphorylation of NKCC2 and KCC3 regulates their plasma membrane expression levels. We identified that spleen tyrosine kinase (SYK) phosphorylates a specific N-terminal tyrosine residue in each cotransporter. Experimental depletion of endogenous SYK or pharmacological inhibition of its kinase activity increased the abundance of NKCC2 at the plasma membrane of human embryonic kidney cells. In contrast, overexpression of a constitutively active SYK mutant decreased NKCC2 membrane abundance. Intriguingly, the same experimental approaches revealed the opposite effect on KCC3 abundance at the plasma membrane, compatible with the known antagonistic roles of NKCC and KCC cotransporters in cell volume regulation. Thus, we identified a novel pathway modulating the cell surface expression of NKCC2 and KCC3 and show that this same pathway has opposite functional outcomes for these two cotransporters. The findings have several biomedical implications considering the role of these cotransporters in regulating blood pressure and cell volume. 

Retracted: No

PubPeer comments:
 Figure 2A. Much more similar than expected.  , 
 Figures 2A and 2C. Much more similar than expected.  , 
 2021 erratum. https: www.sciencedirect.com science article pii S0003986121000643?via%3Dihub The authors regret the poor image quality detected after publication in 2 Western blot panels from Figure 2 and have replaced the corresponding images. The authors would like to apologize for any inconvenience caused. ,