Dominant Functional Role of the Novel Phosphorylation Site S811 in the Human Renal NaCl Cotransporter

FASEB J. 2018 Aug;32(8):4482-4493. doi: 10.1096/fj.201701047R. Epub 2018 Mar 16.


The NaCl cotransporter (NCC) is essential for electrolyte homeostasis and control of blood pressure. The human SLC12A3 gene, which encodes NCC, gives rise to 3 isoforms, of which only the shortest isoform [NaCl cotransporter isoform 3 (NCC3)] has been studied extensively. All NCC isoforms share key phosphorylation sites at T55 and T60 that are essential mediators of NCC function. Recently, a novel phosphorylation site at S811 was identified in isoforms 1 and 2 [NaCl cotransporter splice variant (NCCSV)], which are only present in humans and higher primates. The aim of the current study, therefore, is to investigate the role of S811 phosphorylation in the regulation of NCC by a combination of biochemical and fluorescent microscopy analyses. We demonstrate that hypotonic low-chloride buffer increases S811 phosphorylation, whereas phosphorylation-deficient S811A mutant hinders phosphorylation at T55 and T60 in NCCSV and NCC3. NCCSV S811A impairs NCC3 activity in a dominant-negative fashion, although it does not affect plasma membrane abundance. This effect may be explained by the heterodimerization of NCCSV with NCC3. Taken together, our study highlights the dominant-negative effect of NCCSV on T55 and T60 phosphorylation and NCC activity. Here, we reveal a new function of NCCSV in humans that broadens the understanding on NCC regulation in blood pressure control.-Tutakhel, O. A. Z., Bianchi, F., Smits, D. A., Bindels, R. J. M., Hoenderop, J. G. J., van der Wijst, J. Dominant functional role of the novel phosphorylation site S811 in the human renal NaCl cotransporter.

Keywords: NCC; hypertension; kidney; splice variant.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Blood Pressure / physiology
  • Cell Line
  • HEK293 Cells
  • Humans
  • Kidney / metabolism*
  • Phosphorylation / physiology*
  • Protein Isoforms / metabolism
  • Protein-Serine-Threonine Kinases / metabolism*
  • Solute Carrier Family 12, Member 3 / metabolism*


  • Protein Isoforms
  • Solute Carrier Family 12, Member 3
  • Protein-Serine-Threonine Kinases