Down-regulation of sphingosine kinase-1 by DNA damage: Dependence on proteases and p53

Tarek A. Taha, Walid Osta, Lina Kozhaya, Jacek Bielawski, Korey R. Johnson, William E. Gillanders, Ghassan S. Dbaibo, Yusuf A. Hannun, Lina M. Obeid

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121 Scopus citations


Sphingosine kinase 1 (SK1), a key enzyme in sphingosine 1-phosphate (S1P) synthesis, regulates various aspects of cell behavior, including cell survival and proliferation. DNA damaging anti-neoplastic agents have been shown to induce p53, ceramide levels, and apoptosis; however, the effects of anti-neoplastic agents on SK have not been assessed. In this study, we investigated the effects of a DNA damaging agent, actinomycin D (Act D), on the function of sphingosine kinase (SK1). Act D caused a reduction in the protein levels of SK1, as indicated by Western blot analysis, with a concomitant decrease in SK activity. The down-regulation was posttranscriptional, because the mRNA levels of SK1 remained unchanged. Similar decreases in SK1 protein were observed with other DNA damaging agents such as doxorubicin, etoposide, and γ-irradiation. ZVAD, the pancaspase inhibitor, and Bcl-2 annulled the effect of Act D on SK1, demonstrating a role for cysteine proteases downstream of Bcl-2 in the down-regulation of SKI. Inhibition of caspases 3, 6, 7, and 9 only partially reversed Act D-induced SK1 loss. Inhibition of cathepsin B, a lysosomal protease, produced a significant reversal of SK1 decline by Act D, suggesting that a multitude of ZVAD-sensitive cysteine proteases downstream of Bcl-2 mediated the SK1 decrease. When p53 up-regulation after Act D treatment was inhibited, SK1 down-regulation was rescued, demonstrating p53 dependence of SK1 modulation. Treatment of cells with S1P, the product of SK1, partially inhibited Act D-induced cell death, raising the possibility that a decrease in SK1 may be in part necessary for cell death to occur. Furthermore, the knockdown of SK1 by small interfering RNA in MCF-7 cells resulted in a significant reduction in cell viability. These studies demonstrate that SK1 is down-regulated by genotoxic stress, and that basal SK1 function may be necessary for the maintenance of tumor cell growth.

Original languageEnglish
Pages (from-to)20546-20554
Number of pages9
JournalJournal of Biological Chemistry
Issue number19
StatePublished - May 7 2004


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