title: Protein phosphatase 1-dependent dephosphorylation of Akt is the prime signaling event in sphingosine-induced apoptosis in Jurkat cells.
abstract: Sphingosine (SPH) is an important bioactive lipid involved in mediating a variety of cell functions including apoptosis. However, the signaling mechanism of SPH-induced apoptosis remains unclear. We have investigated whether SPH inhibits survival signaling in cells by inhibiting Akt kinase activity. This study demonstrates that treatment of Jurkat cells with SPH leads to Akt dephosphorylation as early as 15  min, and the cells undergo apoptosis after 6  h. This Akt dephosphorylation is not mediated through deactivation of upstream kinases, since SPH does not inhibit the upstream phosphoinositide-dependent kinase 1 (PDK1) phosphorylation. Rather, sensitivity to the Ser/Thr protein phosphatase inhibitors (calyculin A, phosphatidic acid, tautomycin, and okadaic acid) indicates an important role for protein phosphatase 1 (PP1) in this process. In vitro phosphatase assay, using Akt immunoprecipitate following treatment with SPH, reveals an increase in Akt-PP1 association as determined by immunoprecipitation analysis. Moreover, SPH-induced dephosphorylation of Akt at Ser(473) subsequently leads to the activation of GSK-3β, caspase 3, PARP cleavage, and ultimately apoptosis. Pre-treatment with caspase 3 inhibitor z-VAD-fmk and Ser/Thr phosphatase inhibitor abrogates the effect of SPH on facilitating apoptosis. Altogether, these results demonstrate that PP1-mediated inhibition of the key anti-apoptotic protein, Akt, plays an important role in SPH-mediated apoptosis in Jurkat cells.
authors.affiliation: Cell Signaling Laboratory, Department of Biochemistry, Faculty of Medicine and Health Sciences, UAE University P.O. Box 17666, Al Ain, UAE.
authors.forename: Faisal
authors.initials: F
authors.lastname: Thayyullathil
authors.forename: Shahanas
authors.initials: S
authors.lastname: Chathoth
authors.forename: Allen
authors.initials: A
authors.lastname: Shahin
authors.forename: Jaleel
authors.initials: J
authors.lastname: Kizhakkayil
authors.forename: Abdulkader
authors.initials: A
authors.lastname: Hago
authors.forename: Mahendra
authors.initials: M
authors.lastname: Patel
authors.forename: Sehamuddin
authors.initials: S
authors.lastname: Galadari
mesh_terms: D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D047468:Immunoprecipitation; D019169:Jurkat Cells; D008387:Marine Toxins; D019319:Okadaic Acid / Q000494:pharmacology; D010080:Oxazoles / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D054645:Protein Phosphatase 1 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D011714:Pyrans / Q000494:pharmacology; D012694:Serine / Q000378:metabolism; D015398:Signal Transduction; D013110:Sphingosine / Q000494:pharmacology*; D013141:Spiro Compounds / Q000494:pharmacology
chemical_list: D000590:Amino Acid Chloromethyl Ketones; D004791:Enzyme Inhibitors; D008387:Marine Toxins; D010080:Oxazoles; D011714:Pyrans; D013141:Spiro Compounds; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; C053079:tautomycin; D019319:Okadaic Acid; D012694:Serine; C059041:calyculin A; D011065:Poly(ADP-ribose) Polymerases; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; D051057:Proto-Oncogene Proteins c-akt; D038362:Glycogen Synthase Kinase 3; D054645:Protein Phosphatase 1; D053148:Caspase 3; D013110:Sphingosine