title: JS-K, a nitric oxide-releasing prodrug, modulates ß-catenin/TCF signaling in leukemic Jurkat cells: evidence of an S-nitrosylated mechanism.
abstract: β-Catenin is a central player of the Wnt signaling pathway that regulates cell-cell adhesion and may promote leukemia cell proliferation. We examined whether JS-K, an NO-donating prodrug, modulates the Wnt/β-catenin/TCF-4 signaling pathway in Jurkat T-Acute Lymphoblastic Leukemia cells. JS-K inhibited Jurkat T cell growth in a concentration and time-dependent manner. The IC(50)s for cell growth inhibition were 14±0.7 and 9±1.2μM at 24 and 48h, respectively. Treatment of the cells with JS-K for 24h, caused a dose-dependent increase in apoptosis from 16±3.3% at 10μM to 74.8±2% at 100μM and a decrease in proliferation. This growth inhibition was also due, in part, to alterations in the different phases of the cell cycle. JS-K exhibited a dose-dependent cytotoxicity as measured by LDH release at 24h. However, between 2 and 8h, LDH release was less than 20% for any indicated JS-K concentration. The β-catenin/TCF-4 transcriptional inhibitory activity was reduced by 32±8, 63±5, and 93±2% at 2, 10, and 25μM JS-K, respectively, based on luciferase reporter assays. JS-K reduced nuclear β-catenin and cyclin D1 protein levels, but cytosolic β-catenin expression did not change. Based on a time-course assay of S-nitrosylation of proteins by a biotin switch assay, S-nitrsolyation of nuclear β-catenin was determined to precede its degradation. A comparison of the S-nitrosylated nuclear β-catenin to the total nuclear β-catenin showed that β-catenin protein levels were degraded at 24h, while S-nitrosylation of β-catenin occurred earlier at 0-6h. The NO scavenger PTIO abrogated the JS-K mediated degradation of β-catenin demonstrating the need for NO.
authors.affiliation: Department of Physiology and Pharmacology, City University of New York Medical School, 138th Street and Convent Avenue, New York, NY 10031, United States.
authors.forename: Niharika
authors.initials: N
authors.lastname: Nath
authors.forename: Mitali
authors.initials: M
authors.lastname: Chattopadhyay
authors.forename: Liliya
authors.initials: L
authors.lastname: Pospishil
authors.forename: Lucyna Z
authors.initials: LZ
authors.lastname: Cieciura
authors.forename: Satindra
authors.initials: S
authors.lastname: Goswami
authors.forename: Ravinder
authors.initials: R
authors.lastname: Kodela
authors.forename: Joseph E
authors.initials: JE
authors.lastname: Saavedra
authors.forename: Larry K
authors.initials: LK
authors.lastname: Keefer
authors.forename: Khosrow
authors.initials: K
authors.lastname: Kashfi
mesh_terms: D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D001391:Azo Compounds / Q000494:pharmacology*; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors / Q000502:physiology*; D006131:Growth Inhibitors / Q000494:pharmacology; D006801:Humans; D019169:Jurkat Cells; D009569:Nitric Oxide / Q000378:metabolism* / Q000502:physiology; D009603:Nitroso Compounds / Q000494:pharmacology; D010879:Piperazines / Q000494:pharmacology*; D054218:Precursor T-Cell Lymphoblastic Leukemia-Lymphoma / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D011355:Prodrugs / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D000073940:Transcription Factor 4; D014157:Transcription Factors / Q000502:physiology*; D051153:Wnt Proteins / Q000502:physiology; D051176:beta Catenin / Q000502:physiology*
chemical_list: D001391:Azo Compounds; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D006131:Growth Inhibitors; D009603:Nitroso Compounds; C474958:O(2)-(2,4-dinitrophenyl) 1-((4-ethoxycarbonyl)piperazin-1-yl)diazen-1-ium-1,2-diolate; D010879:Piperazines; D011355:Prodrugs; C532096:TCF4 protein, human; D000073940:Transcription Factor 4; D014157:Transcription Factors; D051153:Wnt Proteins; D051176:beta Catenin; D009569:Nitric Oxide
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