title: P53-dependent upregulation of neutral sphingomyelinase-2: role in doxorubicin-induced growth arrest.
abstract: Neutral sphingomyelinase-2 (nSMase2) is a ceramide-generating enzyme that has been implicated in growth arrest, apoptosis and exosome secretion. Although previous studies have reported transcriptional upregulation of nSMase2 in response to daunorubicin, through Sp1 and Sp3 transcription factors, the role of the DNA damage pathway in regulating nSMase2 remains unclear. In this study, we show that doxorubicin induces a dose-dependent induction of nSMase2 mRNA and protein with concomitant increases in nSMase activity and ceramide levels. Upregulation of nSMase2 was dependent on ATR, Chk1 and p53, thus placing it downstream of the DNA damage pathway. Moreover, overexpression of p53 was sufficient to transcriptionally induce nSMase2, without the need for DNA damage. DNA-binding mutants as well as acetylation mutants of p53 were unable to induce nSMase2, suggesting a role of nSMase2 in growth arrest. Moreover, knockdown of nSMase2 prevented doxorubicin-induced growth arrest. Finally, p53-induced nSMase2 upregulation appears to occur via a novel transcription start site upstream of exon 3. These results identify nSMase2 as a novel p53 target gene, regulated by the DNA damage pathway to induce cell growth arrest.
authors.affiliation: Department of Medicine, Stony Brook University, Health Science Center, Stony Brook, NY 11794-8430, USA.
authors.forename: A A
authors.initials: AA
authors.lastname: Shamseddine
authors.affiliation: Department of Medicine, Stony Brook University, Health Science Center, Stony Brook, NY 11794-8430, USA.
authors.forename: C J
authors.initials: CJ
authors.lastname: Clarke
authors.affiliation: Department of Medicine, Stony Brook University, Health Science Center, Stony Brook, NY 11794-8430, USA.
authors.forename: B
authors.initials: B
authors.lastname: Carroll
authors.affiliation: Department of Medicine, Stony Brook University, Health Science Center, Stony Brook, NY 11794-8430, USA.
authors.forename: M V
authors.initials: MV
authors.lastname: Airola
authors.affiliation: Department of Medicine, Stony Brook University, Health Science Center, Stony Brook, NY 11794-8430, USA.
authors.forename: S
authors.initials: S
authors.lastname: Mohammed
authors.affiliation: Stony Brook University Cancer Center, Stony Brook, NY 11794-8430, USA.
authors.forename: A
authors.initials: A
authors.lastname: Rella
authors.affiliation: Department of Medicine, Stony Brook University, Health Science Center, Stony Brook, NY 11794-8430, USA.|Department of Microbiology and Immunology at Stony Brook University, Stony Brook, NY 11794-8430, USA.
authors.forename: L M
authors.initials: LM
authors.lastname: Obeid
authors.affiliation: Department of Medicine, Stony Brook University, Health Science Center, Stony Brook, NY 11794-8430, USA.|Stony Brook University Cancer Center, Stony Brook, NY 11794-8430, USA.|The Northport Veterans Affairs Hospital, Northport, NY 11768, USA.
authors.forename: Y A
authors.initials: YA
authors.lastname: Hannun
mesh_terms: D004249:DNA Damage; D004317:Doxorubicin / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects; D055785:Gene Knockdown Techniques; D006801:Humans; D061986:MCF-7 Cells; D012333:RNA, Messenger / Q000187:drug effects; D013108:Sphingomyelin Phosphodiesterase / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D015854:Up-Regulation
chemical_list: D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; D004317:Doxorubicin; C491484:SMPD3 protein, human; D013108:Sphingomyelin Phosphodiesterase