Plasma membrane clustering of system y+ (CAT-1) amino acid transporter as detected by immunohistochemistry.
Transport of cationic amino acids in fully differentiated mammalian cells is mediated primarily by system y1+ [cationic amino acid transporter (CAT)-1 gene product]. Antibodies, prepared against synthetic peptide sequences predicted to be extracellular loops of the CAT-1 transporter protein, detected the transporter on the surface of cultured cells. In human fibroblasts, porcine pulmonary artery endothelial cells, and cultured rat hepatoma cells, the CAT-1 transporter protein was clustered in an apparent random pattern throughout the plasma membrane. In contrast, labeling of the fibroblasts with antibodies against the epidermal growth factor receptor or the GLUT-1 glucose transporter demonstrated a uniform staining pattern covering the entire cell surface. The CAT-1 antibody labeling was specific, as demonstrated by peptide inhibition and the lack of staining by preimmune serum. Furthermore, hepatocytes did not exhibit specific antibody binding consistent with the lack of system y1+ activity. Disruption of the microtubule assembly resulted in a reversible loss of the CAT-1 transporter clusters and a more generalized labeling of the cell body. The data demonstrate the existence of microdomains within the plasma membrane that contain the CAT-1 transporter protein.
Woodard
M H
MH
Department of Medicine, College of Medicine, University of Florida, Gainesville 32610-0245.
Dunn
W A
WA
Laine
R O
RO
Malandro
M
M
McMahon
R
R
Simell
O
O
Block
E R
ER
Kilberg
M S
MS
D000595:Amino Acid Sequence; D026923:Amino Acid Transport Systems, Basic; D000596:Amino Acids / Q000378:metabolism*; D000818:Animals; D000906:Antibodies; D001692:Biological Transport; D002352:Carrier Proteins / Q000032:analysis* / Q000096:biosynthesis; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism; D066246:ErbB Receptors / Q000032:analysis / Q000096:biosynthesis; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D051272:Glucose Transporter Type 1; D006801:Humans; D008114:Liver Neoplasms, Experimental / Q000378:metabolism; D008565:Membrane Proteins / Q000032:analysis* / Q000096:biosynthesis; D051379:Mice; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins / Q000032:analysis / Q000096:biosynthesis; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology; D011651:Pulmonary Artery; D051381:Rats; D013552:Swine
D026923:Amino Acid Transport Systems, Basic; D000596:Amino Acids; D000906:Antibodies; D002352:Carrier Proteins; D051272:Glucose Transporter Type 1; D008565:Membrane Proteins; D009004:Monosaccharide Transport Proteins; D010455:Peptides; C495677:SLC2A1 protein, human; C495678:Slc2a1 protein, mouse; C495679:Slc2a1 protein, rat; D066246:ErbB Receptors