Modulatory roles of NHERF1 and NHERF2 in cell surface expression of the glutamate transporter GLAST

Biochem Biophys Res Commun. 2013 Jan 11;430(2):839-45. doi: 10.1016/j.bbrc.2012.11.059. Epub 2012 Nov 27.


The PDZ (PSD-95/Drosophila discs-large protein/zonula occludens protein) domain-containing proteins Na(+)/H(+) exchanger regulatory factor 1 (NHERF1) and NHERF2 interact with the glutamate transporter GLAST. To characterize the roles of these NHERF proteins in the plasma membrane targeting of GLAST, we examined the interaction of green fluorescent protein (EGFP)-tagged GLAST with epitope-tagged NHERF proteins in human embryonic kidney (HEK) 293T cells. Co-expression of either NHERF protein increased the cell surface expression of EGFP-GLAST. Deletion of the C-terminal PDZ domain-binding motif caused an increase in EGFP-GLAST with immature endoglycosidase H-sensitive N-linked oligosaccharides, suggesting impaired exit of EGFP-GLAST from the endoplasmic reticulum (ER). Immunoprecipitation experiments revealed that NHERF1 predominantly bound EGFP-GLAST containing immature N-glycans, whereas NHERF2 co-precipitated EGFP-GLAST with mature N-glycans. Expression of a dominant-negative mutant of the GTPase Sar1 increased the interaction of EGFP-GLAST with NHERF1 in the ER. By contrast, immunofluorescence microscopy showed that NHERF2 co-localized with EGFP-GLAST in ER-Golgi intermediate compartments (ERGICs), at the plasma membrane and in early endosomes, but not in the ER. These results suggest that NHERF1 interacts with GLAST during ER export, while NHERF2 interacts with GLAST in the secretory pathway from the ERGIC to the plasma membrane, thereby modulating the cell surface expression of GLAST.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Membrane / metabolism*
  • Dogs
  • Endoplasmic Reticulum / metabolism
  • Excitatory Amino Acid Transporter 1 / genetics
  • Excitatory Amino Acid Transporter 1 / metabolism*
  • Golgi Apparatus / metabolism
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • HEK293 Cells
  • Humans
  • Monomeric GTP-Binding Proteins / genetics
  • Monomeric GTP-Binding Proteins / metabolism
  • PDZ Domains / genetics
  • Phosphoproteins / genetics
  • Phosphoproteins / metabolism*
  • Sodium-Hydrogen Exchangers / genetics
  • Sodium-Hydrogen Exchangers / metabolism*


  • Excitatory Amino Acid Transporter 1
  • Phosphoproteins
  • SLC1A3 protein, human
  • Sodium-Hydrogen Exchangers
  • enhanced green fluorescent protein
  • sodium-hydrogen exchanger regulatory factor
  • Green Fluorescent Proteins
  • Monomeric GTP-Binding Proteins