Impact of Hybrid and Complex N-Glycans on Cell Surface Targeting of the Endogenous Chloride Cotransporter Slc12a2

International Journal of Cell Biology
Richa SinghMauricio Di Fulvio

Abstract

The Na(+)K(+)2Cl(-) cotransporter-1 (Slc12a2, NKCC1) is widely distributed and involved in cell volume/ion regulation. Functional NKCC1 locates in the plasma membrane of all cells studied, particularly in the basolateral membrane of most polarized cells. Although the mechanisms involved in plasma membrane sorting of NKCC1 are poorly understood, it is assumed that N-glycosylation is necessary. Here, we characterize expression, N-glycosylation, and distribution of NKCC1 in COS7 cells. We show that ~25% of NKCC1 is complex N-glycosylated whereas the rest of it corresponds to core/high-mannose and hybrid-type N-glycosylated forms. Further, ~10% of NKCC1 reaches the plasma membrane, mostly as core/high-mannose type, whereas ~90% of NKCC1 is distributed in defined intracellular compartments. In addition, inhibition of the first step of N-glycan biosynthesis with tunicamycin decreases total and plasma membrane located NKCC1 resulting in almost undetectable cotransport function. Moreover, inhibition of N-glycan maturation with swainsonine or kifunensine increased core/hybrid-type NKCC1 expression but eliminated plasma membrane complex N-glycosylated NKCC1 and transport function. Together, these results suggest that (i) NKCC1 is deliver...Continue Reading

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Citations

Sep 25, 2015·The Journal of Endocrinology·Saeed AlshahraniMauricio Di Fulvio
Jan 20, 2017·Frontiers in Cell and Developmental Biology·Richa SinghMauricio Di Fulvio
Aug 1, 2019·American Journal of Physiology. Cell Physiology·Lisa KellyMauricio Di Fulvio
May 13, 2017·Scientific Reports·Shams KursanMauricio Di Fulvio
Dec 15, 2020·ELife·Dana M TalsnessClement Y Chow
Jun 6, 2021·Cellular and Molecular Life Sciences : CMLS·Yevgen ZolotarovJeroen H F de Baaij
Aug 8, 2021·International Journal of Molecular Sciences·Amanda ChantziouPanayiotis A Theodoropoulos

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Methods Mentioned

BETA
transfection
PCR
electrophoresis
GTPase

Software Mentioned

Image Lab Quantitative
Graphpad Prism
Metamorph
ImageJ
Geneious
MetaVue

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