Long-term effects of rapamycin treatment on insulin mediated phosphorylation of Akt/PKB and glycogen synthase activity.
Protein kinase B (Akt/PKB) is a Ser/Thr kinase that is involved in the regulation of cell proliferation/survival through mammalian target of rapamycin (mTOR) and the regulation of glycogen metabolism through glycogen synthase kinase 3beta (GSK-3beta) and glycogen synthase (GS). Rapamycin is an inhibitor of mTOR. The objective of this study was to investigate the effects of rapamycin pretreatment on the insulin mediated phosphorylation of Akt/PKB phosphorylation and GS activity in parental HepG2 and HepG2 cells with overexpression of constitutively active Akt1/PKB-alpha (HepG2-CA-Akt/PKB). Rapamycin pretreatment resulted in a decrease (20-30%) in the insulin mediated phosphorylation of Akt1 (Ser 473) in parental HepG2 cells but showed an upregulation of phosphorylation in HepG2-CA-Akt/PKB cells. Rictor levels were decreased (20-50%) in parental HepG2 cells but were not significantly altered in the HepG2-CA-Akt/PKB cells. Furthermore, rictor knockdown decreased the phosphorylation of Akt (Ser 473) by 40-60% upon rapamycin pretreatment. GS activity followed similar trends as that of phosphorylated Akt and so with rictor levels in these cells pretreated with rapamycin; parental HepG2 cells showed a decrease in GS activity, whereas as HepG2-CA-Akt/PKB cells showed an increase in GS activity. The changes in the levels of phosphorylated Akt/PKB (Ser 473) correlated with GS and protein phoshatase-1 activity.
Varma
Shailly
S
Department of Biochemistry, College of Medicine, University of Saskatchewan, Saskatoon, SK Canada.
Shrivastav
Anuraag
A
Changela
Sheena
S
Khandelwal
Ramji L
RL
D000730:Androstadienes / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism; D002460:Cell Line; D006006:Glycogen Synthase / Q000378:metabolism*; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D000076226:Rapamycin-Insensitive Companion of mTOR Protein; D015398:Signal Transduction / Q000187:drug effects; D020123:Sirolimus / Q000494:pharmacology*; D013997:Time Factors; D000077191:Wortmannin
D000730:Androstadienes; D002352:Carrier Proteins; D007328:Insulin; C492571:RICTOR protein, human; D000076226:Rapamycin-Insensitive Companion of mTOR Protein; D006006:Glycogen Synthase; D051057:Proto-Oncogene Proteins c-akt; D010749:Phosphoprotein Phosphatases; D020123:Sirolimus; D000077191:Wortmannin