title: Alkaline ceramidase 2 and its bioactive product sphingosine are novel regulators of the DNA damage response.
abstract: Human cells respond to DNA damage by elevating sphingosine, a bioactive sphingolipid that induces programmed cell death (PCD) in response to various forms of stress, but its regulation and role in the DNA damage response remain obscure. Herein we demonstrate that DNA damage increases sphingosine levels in tumor cells by upregulating alkaline ceramidase 2 (ACER2) and that the upregulation of the ACER2/sphingosine pathway induces PCD in response to DNA damage by increasing the production of reactive oxygen species (ROS). Treatment with the DNA damaging agent doxorubicin increased both ACER2 expression and sphingosine levels in HCT116 cells in a dose-dependent manner. ACER2 overexpression increased sphingosine in HeLa cells whereas knocking down ACER2 inhibited the doxorubicin-induced increase in sphingosine in HCT116 cells, suggesting that DNA damage elevates sphingosine by upregulating ACER2. Knocking down ACER2 inhibited an increase in the apoptotic and necrotic cell population and the cleavage of poly ADP ribose polymerase (PARP) in HCT116 cells in response to doxorubicin as well as doxorubicin-induced release of lactate dehydrogenase (LDH) from these cells. Similar to treatment with doxorubicin, ACER2 overexpression induced an increase in the apoptotic and necrotic cell population and PARP cleavage in HeLa cells and LDH release from cells, suggesting that ACER2 upregulation mediates PCD in response to DNA damage through sphingosine. Mechanistic studies demonstrated that the upregulation of the ACER2/sphingosine pathway induces PCD by increasing ROS levels. Taken together, these results suggest that the ACER2/sphingosine pathway mediates PCD in response to DNA damage through ROS production.
authors.affiliation: Department of Medicine, State University of New York at Stony Brook, Stony Brook, NY 11794, USA.|Stony Brook Cancer Center, State University of New York at Stony Brook, Stony Brook, NY 11794, USA.
authors.forename: Ruijuan
authors.initials: R
authors.lastname: Xu
authors.affiliation: Department of Medicine, State University of New York at Stony Brook, Stony Brook, NY 11794, USA.|Stony Brook Cancer Center, State University of New York at Stony Brook, Stony Brook, NY 11794, USA.
authors.forename: Kai
authors.initials: K
authors.lastname: Wang
authors.affiliation: Lipidomics Core Facility, State University of New York at Stony Brook, Stony Brook, NY 11794, USA.
authors.forename: Izolda
authors.initials: I
authors.lastname: Mileva
authors.affiliation: Department of Medicine, State University of New York at Stony Brook, Stony Brook, NY 11794, USA.|Stony Brook Cancer Center, State University of New York at Stony Brook, Stony Brook, NY 11794, USA.
authors.forename: Yusuf A
authors.initials: YA
authors.lastname: Hannun
authors.affiliation: Department of Medicine, State University of New York at Stony Brook, Stony Brook, NY 11794, USA.|Stony Brook Cancer Center, State University of New York at Stony Brook, Stony Brook, NY 11794, USA.|Ralph H. Johnson Veterans Administration Hospital, Stony Brook, NY 11794, USA.
authors.forename: Lina M
authors.initials: LM
authors.lastname: Obeid
authors.affiliation: Department of Medicine, State University of New York at Stony Brook, Stony Brook, NY 11794, USA.|Stony Brook Cancer Center, State University of New York at Stony Brook, Stony Brook, NY 11794, USA.
authors.forename: Cungui
authors.initials: C
authors.lastname: Mao
mesh_terms: D055574:Alkaline Ceramidase / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D045744:Cell Line, Tumor; D004249:DNA Damage; D004317:Doxorubicin / Q000494:pharmacology; D045325:HCT116 Cells; D019073:HT29 Cells; D006367:HeLa Cells; D056945:Hep G2 Cells; D006801:Humans; D017382:Reactive Oxygen Species / Q000378:metabolism; D013110:Sphingosine / Q000378:metabolism*; D015854:Up-Regulation
chemical_list: D017382:Reactive Oxygen Species; D004317:Doxorubicin; C526755:ACER2 protein, human; D055574:Alkaline Ceramidase; D013110:Sphingosine
keywords: Golgi; ceramide; p53; programmed cell death; reactive oxygen species