Differential regulation of the lysosomal, Ca2+-dependent and ubiquitin/proteasome-dependent proteolytic pathways in fast-twitch and slow-twitch rat muscle following hyperinsulinaemia.
In order to characterize the poorly defined mechanisms that account for the anti-proteolytic effects of insulin in skeletal muscle, we investigated in rats the effects of a 3 h systemic euglycaemic hyperinsulinaemic clamp on lysosomal, Ca(2+)-dependent proteolysis, and on ubiquitin/proteasome-dependent proteolysis. Proteolysis was measured in incubated fast-twitch mixed-fibre extensor digitorum longus (EDL) and slow-twitch red-fibre soleus muscles harvested at the end of insulin infusion. Insulin inhibited proteolysis (P<0.05) in both muscles. This anti-proteolytic effect disappeared in the presence of inhibitors of the lysosomal/Ca(2+)-dependent proteolytic pathways in the soleus, but not in the EDL, where only the proteasome inhibitor MG 132 (benzyloxycarbonyl-leucyl-leucyl-leucinal) was effective. Furthermore, insulin depressed ubiquitin mRNA levels in the mixed-fibre tibialis anterior, but not in the red-fibre diaphragm muscle, suggesting that insulin inhibits ubiquitin/proteasome-dependent proteolysis in mixed-fibre muscles only. However, depressed ubiquitin mRNA levels in such muscles were not associated with significant decreases in the amount of ubiquitin conjugates, or in mRNA levels or protein content for the 14 kDa ubiquitin-conjugating enzyme E2 and 20 S proteasome subunits. Thus alternative, as yet unidentified, mechanisms are likely to contribute to inhibit the ubiquitin/proteasome system in mixed-fibre muscles.
Larbaud
D
D
Human Nutrition Research Center of Clermont-Ferrand, and Institut National de la Recherche Agronomique, Nutrition and Protein Metabolism Unit, 63122 Ceyrat, France.
Balage
M
M
Taillandier
D
D
Combaret
L
L
Grizard
J
J
Attaix
D
D
D000818:Animals; D001786:Blood Glucose / Q000378:metabolism; D002118:Calcium / Q000502:physiology; D046508:Culture Techniques; D003546:Cysteine Endopeptidases / Q000378:metabolism; D006946:Hyperinsulinism / Q000097:blood / Q000378:metabolism*; D007328:Insulin / Q000097:blood / Q000502:physiology; D008099:Liver / Q000201:enzymology; D008247:Lysosomes / Q000378:metabolism; D008297:Male; D009097:Multienzyme Complexes / Q000378:metabolism; D009124:Muscle Proteins / Q000378:metabolism*; D018482:Muscle, Skeletal / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D025801:Ubiquitin / Q000378:metabolism
D001786:Blood Glucose; D007328:Insulin; D009097:Multienzyme Complexes; D009124:Muscle Proteins; D012333:RNA, Messenger; D025801:Ubiquitin; C069767:phosphoenolpyruvate carboxylase kinase; D017346:Protein Serine-Threonine Kinases; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D002118:Calcium