The T-tubule is a cell-surface target for insulin-regulated recycling of membrane proteins in skeletal muscle.
(1) In this study we have determined the distribution of various membrane proteins involved in insulin-activated glucose transport in T-tubules and in sarcolemma from rat skeletal muscle. Two independent experimental approaches were used to determine the presence of membrane proteins in T-tubules: (i) the purification of T-tubules free from sarcolemmal membranes by lectin agglutination, and (ii) T-tubule vesicle immunoadsorption. These methods confirmed that T-tubules from rat skeletal muscle were enriched with dihydropyridine receptors and tt28 protein and did not contain the sarcolemmal markers dystrophin or beta 1-integrin. Both types of experiments revealed an abundant content of GLUT4 glucose carriers, insulin receptors and SCAMPs (secretory carrier membrane proteins) in T-tubule membranes. (2) Acute administration in vivo of insulin caused an increased abundance of GLUT4 in T-tubules and sarcolemma. On the contrary, insulin led to a 50% reduction in insulin receptors present in T-tubules and in sarcolemma, demonstrating that insulin-induced insulin receptor internalization affects T-tubules in the muscle fibre. The alteration in the content of GLUT4 and insulin receptors in T-tubules was a consequence of insulin-induced redistribution of these proteins. SCAMPs also redistributed in muscle membranes in response to insulin. They were recruited by insulin from intracellular high-density fractions to intracellular lighter-density fractions and to the cell surface, showing a pattern of insulin-induced cellular redistribution distinct from those of GLUT4 and the insulin receptor. (3) In conclusion, the T-tubule is a cell-surface target for membrane proteins involved in recycling such as SCAMPs or for membrane proteins that acutely redistribute in response to insulin such as GLUT4 or insulin receptors.
Departament de Bioquímica i Fisiologia, Facultat de Biologia, Universitat de Barcelona, Spain.
P
P
Muñoz
M
M
Rosemblatt
X
X
Testar
M
M
Palacín
G
G
Thoidis
P F
PF
Pilch
A
A
Zorzano
D000818:Animals; D015220:Calcium Channels / Q000302:isolation & purification / Q000378:metabolism; D020746:Calcium Channels, L-Type; D002352:Carrier Proteins / Q000032:analysis / Q000378:metabolism*; D002458:Cell Fractionation; D051275:Glucose Transporter Type 4; D007328:Insulin / Q000494:pharmacology*; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism*; D009004:Monosaccharide Transport Proteins / Q000032:analysis / Q000378:metabolism*; D009124:Muscle Proteins; D018482:Muscle, Skeletal / Q000187:drug effects / Q000378:metabolism*; D015388:Organelles / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D011972:Receptor, Insulin / Q000302:isolation & purification / Q000378:metabolism*; D012508:Sarcolemma / Q000378:metabolism*
D015220:Calcium Channels; D020746:Calcium Channels, L-Type; D002352:Carrier Proteins; D051275:Glucose Transporter Type 4; D007328:Insulin; D008565:Membrane Proteins; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; C495694:Slc2a4 protein, rat; D011972:Receptor, Insulin