The GLUT4 glucose transporter and the alpha 2 subunit of the Na+,K(+)-ATPase do not localize to the same intracellular vesicles in rat skeletal muscle.
The GLUT4 glucose transporter and the alpha 2 subunit of the Na+,K(+)-ATPase of rat skeletal muscle are two proteins which redistribute from intracellular membranes to plasma membranes following in vivo insulin stimulation. Here we show that although both proteins co-segregate after subcellular fractionation of unstimulated rat hindlimb muscles, they do not share the same intracellular residence inside the muscle fibre. By immunogold single- and double-labeling on ultrathin muscle cryosections with specific antibodies, the GLUT4 glucose transporter and the Na+,K(+)-ATPase alpha 2 subunit were observed on different vesicular structures within the cell. GLUT4 was detected on subsarcolemmal and perinuclear membranes, and at the junction between myofibrillar A and I bands where triads are localized. The alpha 2 subunit of the Na+,K(+)-ATPase was observed at the plasma membrane and in distinct subsarcolemmal vesicles and intermyofibrillar membranes. Quantitative analysis of double-labeling of GLUT4 and Na+,K(+)-ATPase alpha 2 subunit revealed that less than 6% of the two proteins co-localize in the same continuous vesicular structures. The differential intracellular localization of the two proteins was further confirmed by immunopurification of GLUT4-containing membranes from muscle homogenates, in which the alpha 2 subunit of the Na+,K(+)-ATPase was found only at the same extent as the alpha 1 subunit of the enzyme, a protein exclusively present at the plasma membrane.
Lavoie
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L
Division of Cell Biology, Hospital for Sick Children, Toronto, Ont., Canada.
He
L
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Ramlal
T
T
Ackerley
C
C
Marette
A
A
Klip
A
A
D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry; D029968:Escherichia coli Proteins; D051275:Glucose Transporter Type 4; D006026:Glycoside Hydrolases; D016695:Glycosyltransferases / Q000737:chemistry; D007150:Immunohistochemistry; D007328:Insulin / Q000494:pharmacology; D007425:Intracellular Membranes / Q000378:metabolism*; D008297:Male; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*; D009124:Muscle Proteins / Q000378:metabolism*; D009132:Muscles / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*; D013347:Subcellular Fractions / Q000737:chemistry
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D051275:Glucose Transporter Type 4; D007328:Insulin; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; C495694:Slc2a4 protein, rat; D016695:Glycosyltransferases; D006026:Glycoside Hydrolases; C085428:slt protein, E coli; D000254:Sodium-Potassium-Exchanging ATPase