WNK1 phosphorylation sites in TBC1D1 and TBC1D4 modulate cell surface expression of GLUT1

Andreia F.A. Henriques, Paulo Matos, Ana Sofia Carvalho, Mikel Azkargorta, Felix Elortza, Rune Matthiesen, Peter Jordan

Research output: Contribution to journalArticle

Abstract

Glucose uptake by mammalian cells is a key mechanism to maintain cell and tissue homeostasis and relies mostly on plasma membrane-localized glucose transporter proteins (GLUTs). Two main cellular mechanisms regulate GLUT proteins in the cell: first, expression of GLUT genes is under dynamic transcriptional control and is used by cancer cells to increase glucose availability. Second, GLUT proteins are regulated by membrane traffic from storage vesicles to the plasma membrane (PM). This latter process is triggered by signaling mechanisms and well-studied in the case of insulin-responsive cells, which activate protein kinase AKT to phosphorylate TBC1D4, a RAB-GTPase activating protein involved in membrane traffic regulation. Previously, we identified protein kinase WNK1 as another kinase able to phosphorylate TBC1D4 and regulate the surface expression of the constitutive glucose transporter GLUT1. Here we describe that downregulation of WNK1 through RNA interference in HEK293 cells led to a 2-fold decrease in PM GLUT1 expression, concomitant with a 60% decrease in glucose uptake. By mass spectrometry, we identified serine (S) 704 in TBC1D4 as a WNK1-regulated phosphorylation site, and also S565 in the paralogue TBC1D1. Transfection of the respective phosphomimetic or unphosphorylatable TBC1D mutants into cells revealed that both affected the cell surface abundance of GLUT1. The results reinforce a regulatory role for WNK1 in cell metabolism and have potential impact for the understanding of cancer cell metabolism and therapeutic options in type 2 diabetes.

Original languageEnglish
Article number108223
JournalArchives of Biochemistry and Biophysics
Volume679
DOIs
Publication statusPublished - 15 Jan 2020

Fingerprint

Phosphorylation
Facilitative Glucose Transport Proteins
Cell membranes
Cells
Metabolism
Glucose
Protein Kinases
Tissue homeostasis
Membranes
GTPase-Activating Proteins
Cell Membrane
Medical problems
Serine
Mass spectrometry
Phosphotransferases
Genes
Availability
RNA
Insulin
HEK293 Cells

Keywords

  • Glucose cotransport
  • GLUT1
  • Membrane traffic
  • Protein phosphorylation
  • TBC1D
  • WNK1

Cite this

Henriques, Andreia F.A. ; Matos, Paulo ; Carvalho, Ana Sofia ; Azkargorta, Mikel ; Elortza, Felix ; Matthiesen, Rune ; Jordan, Peter. / WNK1 phosphorylation sites in TBC1D1 and TBC1D4 modulate cell surface expression of GLUT1. In: Archives of Biochemistry and Biophysics. 2020 ; Vol. 679.
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WNK1 phosphorylation sites in TBC1D1 and TBC1D4 modulate cell surface expression of GLUT1. / Henriques, Andreia F.A.; Matos, Paulo; Carvalho, Ana Sofia; Azkargorta, Mikel; Elortza, Felix; Matthiesen, Rune; Jordan, Peter.

In: Archives of Biochemistry and Biophysics, Vol. 679, 108223, 15.01.2020.

Research output: Contribution to journalArticle

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AU - Henriques, Andreia F.A.

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AU - Carvalho, Ana Sofia

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AU - Elortza, Felix

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AU - Jordan, Peter

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KW - Glucose cotransport

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