title: Eicosapentaenoic acid demethylates a single CpG that mediates expression of tumor suppressor CCAAT/enhancer-binding protein delta in U937 leukemia cells.
abstract: Polyunsaturated fatty acids (PUFAs) inhibit proliferation and induce differentiation in leukemia cells. To investigate the molecular mechanisms whereby fatty acids affect these processes, U937 leukemia cells were conditioned with stearic, oleic, linolenic, α-linolenic, arachidonic, eicosapentaenoic, and docosahexaenoic acids. PUFAs affected proliferation; eicosapentaenoic acid (EPA) was the most potent on cell cycle progression. EPA enhanced the expression of the myeloid lineage-specific transcription factors CCAAT/enhancer-binding proteins (C/EBPβ and C/EBPδ), PU.1, and c-Jun, resulting in increased expression of the monocyte lineage-specific target gene, the macrophage colony-stimulating factor receptor. Indeed, it is known that PU.1 and C/EBPs interact with their consensus sequences on a small DNA fragment of macrophage colony-stimulating factor receptor promoter, which is a determinant for expression. We demonstrated that C/EBPβ and C/EBPδ bind the same response element as a heterodimer. We focused on the enhanced expression of C/EBPδ, which has been reported to be a tumor suppressor gene silenced by promoter hypermethylation in U937 cells. After U937 conditioning with EPA and bisulfite sequencing of the -370/-20 CpG island on the C/EBPδ promoter region, we found a site-specific CpG demethylation that was a determinant for the binding activity of Sp1, an essential factor for C/EBPδ gene basal expression. Our results provide evidence for a new role of PUFAs in the regulation of gene expression. Moreover, we demonstrated for the first time that re-expression of the tumor suppressor C/EBPδ is controlled by the methylation state of a site-specific CpG dinucleotide.
authors.affiliation: Department of Experimental Medicine and Biochemical Sciences, University of Perugia, Perugia 06126, Italy.
authors.forename: Veronica
authors.initials: V
authors.lastname: Ceccarelli
authors.forename: Serena
authors.initials: S
authors.lastname: Racanicchi
authors.forename: Maria Paola
authors.initials: MP
authors.lastname: Martelli
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Nocentini
authors.forename: Katia
authors.initials: K
authors.lastname: Fettucciari
authors.forename: Carlo
authors.initials: C
authors.lastname: Riccardi
authors.forename: Pierfrancesco
authors.initials: P
authors.lastname: Marconi
authors.forename: Paolo
authors.initials: P
authors.lastname: Di Nardo
authors.forename: Francesco
authors.initials: F
authors.lastname: Grignani
authors.forename: Luciano
authors.initials: L
authors.lastname: Binaglia
authors.forename: Alba
authors.initials: A
authors.lastname: Vecchini
mesh_terms: D001483:Base Sequence; D051743:CCAAT-Enhancer-Binding Protein-delta / Q000235:genetics / Q000378:metabolism*; D047369:Chromatin Immunoprecipitation; D018899:CpG Islands; D019175:DNA Methylation / Q000187:drug effects*; D017931:DNA Primers; D015118:Eicosapentaenoic Acid / Q000494:pharmacology*; D024202:Electrophoretic Mobility Shift Assay; D005434:Flow Cytometry; D006801:Humans; D009857:Oncogenes; D016133:Polymerase Chain Reaction; D020298:U937 Cells
chemical_list: D017931:DNA Primers; D051743:CCAAT-Enhancer-Binding Protein-delta; D015118:Eicosapentaenoic Acid