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  • 1
    Keywords: Germany ; IN-VIVO ; SUPPORT ; GENE ; GENOME ; PROTEIN ; PROTEINS ; RNA ; TIME ; DNA ; FAMILY ; SUFFICIENT ; STAGE ; Drosophila ; MELANOGASTER ; DNA methylation ; EMBRYO ; FISSION YEAST ; METHYLTRANSFERASE ACTIVITY ; OVEREXPRESSION ; METHYLATION ; CATALYTIC ACTIVITY ; CYTOSINE-5 METHYLTRANSFERASES ; DE-NOVO METHYLATION ; DNA methylation,Drosophila,DNA methyltransferase,Dnmt2,Su(var)3-9 ; DNA methyltransferase ; EMBRYONIC STEM-CELLS ; EMBRYOS ; HISTONE H3 METHYLTRANSFERASE ; HYPERMETHYLATION ; LETHALITY ; MAMMALIAN DEVELOPMENT ; METHYLTRANSFERASE GENE ; SEQUENCE SPECIFICITY
    Abstract: The methylation status of Drosophila DNA has been discussed controversially over a long time. Recent evidence has provided strong support for the existence of 5-methylcytosine in DNA preparations from embryonic stages of fly development. The Drosophila genome contains a single candidate DNA methyltransferase gene that has been termed Dnmt2. This gene belongs to a widely conserved family of putative DNA methyltransferases. However, no catalytic activity has been demonstrated for any Dnmt2-like protein yet. We have now established a protocol for the immunological detection of methylated cytosine in fly embryos. Confocal analysis of immunostained embryos provided direct evidence for the methylation of embryonic DNA. In order to analyse the function of Dnmt2 in DNA methylation, we depleted the protein by RNA interference. Depletion of Dnmt2 had no detectable effect on embryonic development and resulted in a complete loss of DNA methylation. Consistently, overexpression of Dnmt2 from an inducible transgene resulted in significant genomic hypermethylation at CpT and CpA dinucleotides. These results demonstrate that Dnmt2 is both necessary and sufficient for DNA methylation in Drosophila and suggest a novel CpT/A-specific DNA methyltransferase activity for Dnmt2 proteins
    Type of Publication: Journal article published
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  • 2
    Keywords: CELLS ; tumor ; CELL ; Germany ; GENE ; GENES ; PROTEIN ; PROTEINS ; RNA ; transcription ; COMPONENTS ; ACTIVATION ; COMPLEX ; COMPLEXES ; MECHANISM ; TRANSCRIPTION FACTOR ; mechanisms ; BIOLOGY ; MOLECULAR-BIOLOGY ; TRANSCRIPTION FACTORS ; IDENTIFICATION ; CHROMATIN ; Drosophila ; DROSOPHILA-MELANOGASTER ; MELANOGASTER ; PROMOTER ; PROMOTERS ; DNA-BINDING ; DOUBLE-STRANDED-RNA ; REPRESSION ; BINDING PROTEIN ; molecular biology ; molecular ; RE ; INTERFERENCE ; RNA INTERFERENCE ; MOLECULAR-MECHANISMS ; analysis ; SCREEN ; NUCLEAR ; TECHNOLOGY ; ANDROGEN RECEPTOR ; RNAi ; USA ; modification ; CONJUGATION ; SUMO ; CELL BIOLOGY ; GENOME-WIDE ; FACTOR SP3 ; SUMOYLATION
    Abstract: SUMO modification of many transcription factors is linked to transcriptional repression. The molecular mechanisms by which SUMO attachment represses transcription are largely unknown. Here we report a genome-wide RNA interference screen in Drosophila melanogaster cells for components regulating and mediating SUMO-dependent transcriptional repression. Analysis of 〉21,000 double-stranded RNAs (dsRNAs) identified 120 genes whose dsRNA-mediated knockdowns impaired SUMO-dependent transcriptional repression. Several of these genes encode chromatin-associated proteins, including the ATP-dependent chromatin remodeler Mi-2, the D. melanogaster ortholog of the C. elegans protein MEP-1, and the polycomb protein Sfmbt. Knockdown of these proteins did not impair SUMO conjugation, demonstrating that they act downstream of SUMO attachment. Biochemical analyses revealed that MEP1, Mi-2, and Sfmbt interact with each other, bind to SUMO, and are recruited to promoters in a SUMOy-lation-dependent manner. Our results suggest that MEP-1, Mi-2, and Sfmbt are part of a common repression complex established by DNA-bound SUMOmodified transcription factors
    Type of Publication: Journal article published
    PubMed ID: 18374648
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