Heterologously expressed GLT-1 associates in approximately 200-nm protein-lipid islands.
The glutamate transporter GLT-1 from Rattus norvegicus was expressed at high level in baby hamster kidney (BHK-21) cells by the Semliki Forest Virus expression system. We examined the expressed GLT-1 in the plasma membrane and found that the transporter accumulates in detergent-insoluble lipid-protein assemblies. Freeze-fracture, immunogold labeling, and electron microscopy revealed that GLT-1 forms approximately 200-nm protein-rich islands in the plasma membrane. Cholesterol depletion in living cells resulted in a dispersion of the GLT-1 islands, indicating that they are the result of lipid-protein rather than protein-protein interactions. Disruption of GLT-1 islands and dispersion of GLT-1 goes along with a reduction of the glutamate transport activity. Our direct visualization of lipid-protein islands in the plasma membrane of tissue culture cells suggests that the reported clustering of glutamate transporters and their cholesterol-dependent transport activity in cells is likewise connected to their association with cholesterol-rich microdomains in the plasma membrane.
Raunser
Stefan
S
Department of Structural Biology, Max-Planck-Institute of Biophysics, Frankfurt am Main, Germany.
Haase
Winfried
W
Franke
Cornelia
C
Eckert
Gunter P
GP
Müller
Walter E
WE
Kühlbrandt
Werner
W
D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D027342:Excitatory Amino Acid Transporter 2 / Q000235:genetics / Q000378:metabolism*; D007668:Kidney / Q000737:chemistry / Q000378:metabolism*; D008563:Membrane Lipids / Q000737:chemistry / Q000378:metabolism*; D021962:Membrane Microdomains / Q000378:metabolism*; D049329:Nanostructures / Q000737:chemistry / Q000648:ultrastructure; D010316:Particle Size; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism*
D027342:Excitatory Amino Acid Transporter 2; D008563:Membrane Lipids; D011994:Recombinant Proteins