Protein phosphatase 2A regulates apoptosis in intestinal epithelial cells.
Polyamine depletion prevents apoptosis by increasing serine/threonine phosphorylation leading to either inactivation or activation of pro- and anti-apoptotic proteins, respectively. Despite evidence that protein kinases are regulators of apoptosis, a specific role for protein phosphatases in regulating cell survival has not been established. In this study, we show that polyamine depletion inhibits serine/threonine phosphatase 2A (PP2A). Inhibition of PP2A in cells depleted of polyamines correlated well with increased phosphorylation of Bad at Ser112. Bad Ser112 phosphorylation in response to tumor necrosis factor (TNF)-alpha treatment decreased with time in cells grown in control as well as those grown in the presence of alpha-difluoromethylornithine plus putrescine. However, a sustained increase in the levels of Bad Ser112 phosphorylation was maintained in response to TNF-alpha treatment in cells grown in the presence of alpha-difluoromethylornithine. Inhibition of PP2A by okadaic acid and fostriecin or PP2A small interfering RNA transfection significantly decreased TNF-alpha-induced apoptosis in control and polyamine-depleted cells. Inhibition of PP2A by okadaic acid: 1) increased Bad and Bcl-2 phosphorylation at Ser112 and Ser70, respectively; 2) increased ERK activity; 3) prevented JNK activation; 4) prevented cytochrome c release, and activation of caspases-9 and -3 in response to TNF-alpha. Inhibition of MEK1 by U0126 prevented phosphorylation of Bad at Ser112. These results indicate that polyamines regulate PP2A activity, and inhibition of PP2A in response to polyamine depletion increases steady state levels of Bad and Bcl-2 proteins and their phosphorylation and thereby prevents cytochrome c release, caspase-9, and caspase-3 activation.
Department of Physiology, University of Tennessee Health Science Center, Memphis, Tennessee 38163, USA. rray@physio1.utmem.edu
Ramesh M
RM
Ray
Sujoy
S
Bhattacharya
Leonard R
LR
Johnson
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D002352:Carrier Proteins / Q000378:metabolism; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D045304:Cytochromes c / Q000378:metabolism; D000518:Eflornithine / Q000378:metabolism; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000378:metabolism; D004847:Epithelial Cells / Q000166:cytology / Q000502:physiology*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D007413:Intestinal Mucosa / Q000166:cytology*; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D051379:Mice; D019319:Okadaic Acid / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011073:Polyamines / Q000378:metabolism*; D054648:Protein Phosphatase 2; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D051381:Rats; D012694:Serine / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D051025:bcl-Associated Death Protein
C494820:Bad protein, mouse; C494821:Bad protein, rat; D002352:Carrier Proteins; D004791:Enzyme Inhibitors; D011073:Polyamines; D019253:Proto-Oncogene Proteins c-bcl-2; D034741:RNA, Small Interfering; D014409:Tumor Necrosis Factor-alpha; D051025:bcl-Associated Death Protein; D019319:Okadaic Acid; D012694:Serine; D045304:Cytochromes c; D048049:Extracellular Signal-Regulated MAP Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2; C505409:Casp3 protein, mouse; C505410:Casp3 protein, rat; C506434:Casp9 protein, mouse; C506435:Casp9 protein, rat; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases; D000518:Eflornithine