Decreased apoptosis in polyamine depleted IEC-6 cells depends on Akt-mediated NF-kappaB activation but not GSK3beta activity.
The PI3-kinase/Akt pathway promotes cell survival in many different cell types including intestinal epithelial cells. Increased AKT activation in polyamine depleted intestinal epithelial cells correlated well with the decrease in TNF-alpha-induced apoptosis. Increased Akt activation and GSK3beta (Ser 9) phosphorylation without significant effect on Bad (Ser136) phosphorylation indicate that Akt-mediated protection is independent of Bad phosphorylation but may depend on GSK3beta. Pretreatment of polyamine-depleted cells with LY294002 increased caspase-9 and caspase-3 activation and decreased basal levels of GSK-3beta phosphorylation. Inhibition of GSK3beta activity using AR-A014418 or lithium chloride or siRNA-mediated downregulation of its expression had no effect on apoptosis. Inhibition of PI3-kinase and over-expression of dominant negative Akt (DN-AKT), significantly increased apoptosis in polyamine depleted cells. DN-Akt expression reversed the protective effect of polyamine depletion on apoptosis. DN-Akt, as well as the PI3-kinase inhibitors, prevented Akt activation and subsequent translocation of NF-kappaB to the nucleus. Constitutively active Akt (CA-AKT) expression increased resistance to TNF-alpha-induced apoptosis. Constitutively active-Akt expression increased nuclear staining of NF-kappaB. Moreover, polyamine depletion of DN-Akt cells prevented basal and TNF-alpha-induced IkappaBalpha phosphorylation. Prevention of NF-kappaB activation in DN-IkappaBalpha-transfected cells increased apoptosis in control cells and restored it in polyamine-depleted cells to control levels. These data indicate that Akt regulates the mitochondrial pathway, preventing activation of caspase-9 and thereby caspase-3 via NF-kappaB and these effects are independent of GSK-3beta activity.
Bhattacharya
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Department of Physiology, The University of Tennessee Health Science Center, 894 Union Ave., Memphis, Tennessee 38163, USA.
Ray
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RM
Johnson
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D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D053148:Caspase 3 / Q000378:metabolism; D053453:Caspase 9 / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D002867:Chromones / Q000494:pharmacology; D000518:Eflornithine / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D005799:Genes, Dominant; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D020672:I-kappa B Proteins / Q000378:metabolism; D009025:Morpholines / Q000494:pharmacology; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation / Q000187:drug effects; D010768:Phosphoserine / Q000378:metabolism; D011073:Polyamines / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D000077191:Wortmannin; D051025:bcl-Associated Death Protein / Q000378:metabolism
D000730:Androstadienes; D002867:Chromones; D020672:I-kappa B Proteins; D009025:Morpholines; D016328:NF-kappa B; C000606206:Nfkbia protein, rat; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011073:Polyamines; D014409:Tumor Necrosis Factor-alpha; D051025:bcl-Associated Death Protein; D000072000:NF-KappaB Inhibitor alpha; D010768:Phosphoserine; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D000071679:Glycogen Synthase Kinase 3 beta; C000605508:Gsk3b protein, rat; D051057:Proto-Oncogene Proteins c-akt; D038362:Glycogen Synthase Kinase 3; D053148:Caspase 3; D053453:Caspase 9; D000077191:Wortmannin; D000518:Eflornithine