Project description:We investigated the regions that are occupied by deltaNp63 in BxPC-3 and L3.6pl and identification of super enhancers in different pancreatic cancer cell lines. Thereby, we identified a group of 45 super enhancers that are associated with poorer prognosis and are highly dependent on deltaNp63.
Project description:Piwi interacting (pi)RNAs repress diverse transposable elements in the germ cells of metazoans and are essential for fertility in both invertebrates and vertebrates. The precursors of piRNAs are transcribed from distinct genomic regions, the so-called piRNA clusters; however, how piRNA clusters are differentiated from the rest of the genome is not known. To address this question, we studied piRNA biogenesis in two Drosophila virilis strains that show differential ability to generate piRNAs from several genomic regions. We found that active piRNA biogenesis correlates with high levels of histone 3 lysine 9 trimethylation (H3K9me3) over genomic regions that give rise to piRNAs. Furthermore, piRNA biogenesis in the progeny requires the trans-generational inheritance of an epigenetic signal, presumably in form of homologous piRNAs that are generated in the maternal germline and deposited into the oocyte. The inherited piRNAs enhance piRNA biogenesis by installment of H3K9me3 mark on piRNA clusters and by promoting ping-pong processing of homologous transcripts into mature piRNAs. We submitted the resequencing data together with the functional genomic datasets because it was generated with the sole purpose of supporting those. The SRA accession numbers are SRR1536176 and SRR1536175. ChIP-seq against H3K9me3 and Pol2, Total RNA-seq, in Drosophila virilis Strain9 and Strain160 as well as crosses between them
Project description:Translocations producing rearranged versions of the transcription factor DUX4 (DUX4-r) are one of the most frequent causes of B-cell acute lymphoblastic leukemia (B-ALL). DUX4-r retains the DNA binding domain of wt DUX4, but is truncated on the C-terminal transcription activation domain. The precise mechanism through which DUX4-r causes leukemia is unknown and no targeted therapy is currently available. We found that the rearrangement leads to both a loss and a gain of function in DUX4-r. Loss of CBP/EP300 transcriptional co-activators interaction and inability to bind and activate repressed chromatin. Gain of interaction with the transcription factor GTF2I, which redirects DUX4-r toward leukemogenic targets. Importantly, this neomorphic activity exposes an Achilles' heel whereby DUX4-r positive leukemia cells are exquisitely sensitive to GTF2I targeting, which inhibits DUX4-r leukemogenic activity. Our work elucidates the molecular mechanism through which DUX4-r causes leukemia and suggest a possible therapeutic avenue tailored to this B-ALL subtype.
Project description:Small non-coding RNAs that associate with Piwi proteins, called piRNAs, serve as guides for repression of diverse transposable elements in germ cells of Metazoa. In Drosophila, the genomic regions that give rise to piRNAs, the so-called piRNA clusters, are transcribed to generate long precursor molecules that are processed into mature piRNAs. How genomic regions that give rise to piRNA precursor transcripts are differentiated from the rest of the genome and how these transcripts are specifically channeled into the piRNA biogenesis pathway are not known. We found that trans-generationally inherited piRNAs provide the critical trigger for piRNA production from homologous genomic regions in the next generation by two different mechanisms. First, inherited piRNAs enhance processing of homologous transcripts into mature piRNAs by initiating the ping-pong cycle in the cytoplasm. Second, inherited piRNAs induce installment of the H3K9me3 mark on genomic piRNA cluster sequences. The HP1 homolog Rhino binds to the H3K9me3 mark through its chromodomain and is enriched over piRNA clusters. Rhino recruits the piRNA biogenesis factor Cutoff to piRNA clusters and is required for efficient transcription of piRNA precursors. We propose that trans-generationally inherited piRNAs act as an epigenetic memory for identification of substrates for piRNA biogenesis on two levels, by inducing a permissive chromatin environment for piRNA precursor synthesis and by enhancing processing of these precursors. ChIPseq of Rhino and Cutoff in Drosophila melanogaster ovaries The Rhino-BioTAP flies were made by fusing the BioTAP tag (Alekseyenko et al. 2014 )to the C-terminal region of the Rhino gene under the UASp promoter. The Cutoff-EGFP fly line (Nanos-GAL4/UASp-Cutoff-GFP), Cuff^wm25 and Cuff^qq37 were a generous gift from T. Schupbach.
Project description:Aim: To determine Myc binding sites in mouse heart and liver, 4 hours post MycER activation. c-Myc ChIP sequencing performed on chromatin isolated from hearts and livers harvested from wild-type (R26+/+) and R26CAG-c-MycERT2/+ mice 4 hours post administration of (Z)-4-hydroxytamoxifen.
Project description:Aim: To determine RNA Pol II binding sites in mouse heart and liver, 4 hours post MycER activation. Pol II ChIP sequencing performed on chromatin isolated from hearts and livers harvested from wild-type (R26+/+) and R26CAG-c-MycERT2/+ mice 4 hours post administration of (Z)-4-hydroxytamoxifen.
Project description:In mammals, one of the female X chromosomes and all imprinted genes are expressed exclusively from a single allele in somatic cells. To evaluate structural changes associated with allelic silencing, we have applied a recently developed Hi-C assay that uses DNase I for chromatin fragmentation to mouse F1 hybrid systems. Results We find radically different conformations for the two female mouse X chromosomes. The inactive X has two superdomains of frequent intrachromosomal contacts separated by a boundary/hinge region. Comparison with the recently reported bipartite 3D structure of the human inactive X shows that the genomic content of the superdomains differs between species, but part of the hinge is conserved and located near the Dxz4/DXZ4 locus. In mouse, the hinge region also contains a minisatellite Ds-TR, and both Dxz4 and Ds-TR appear to be anchored to the nucleolus. Genes that escape X inactivation do not cluster but are located near the periphery of the 3D structure, as are regions enriched in CTCF or RNA polymerase. Fewer short-range intrachromosomal contacts are detected for the inactive alleles of genes subject to X inactivation, compared to the active alleles and to genes that escape X inactivation. This pattern is also evident for imprinted genes, in which more chromatin contacts are detected for the expressed allele. Conclusions By applying a novel Hi-C method to map allelic chromatin contacts, we discover a specific bipartite organization of the mouse inactive X chromosome that probably plays an important role in maintenance of gene silencing. Examination of Nucleophosmin-binding profiles in male and female mouse livers
Project description:In the Metazoan germline, piwi proteins and associated piwi-interacting RNAs (piRNAs) provide a defense system against the expression of transposable elements. In the cytoplasm, piRNA sequences guide piwi complexes to destroy complementary transposon transcripts by endonucleolytic cleavage. However, some piwi family members are nuclear, raising the possibility of alternative pathways for piRNA-mediated regulation of gene expression. We found that Drosophila Piwi is recruited to chromatin, co-localizing with RNA polymerase II on polytene chromosomes. Knockdown of Piwi in the germline increases expression of transposable elements that are targeted by piRNAs, whereas protein-coding genes remain largely unaffected. De-repression of transposons upon Piwi depletion correlates with increased occupancy of RNA polymerase II on their promoters. Expression of piRNAs that target a reporter construct results in a decrease in Pol II occupancy and an increase in repressive H3K9me3 marks and HP1 on the reporter locus. Our results indicate that Piwi identifies targets complementary to the associated piRNA and induces transcriptional repression by establishing a repressive chromatin state when correct targets are found. Examination of mRNA levels, Pol II occupancy, and chromatin repressive state in D. melanogaster ovaries upon Piwi knockdown and Piwi mutation. This records present the sequencing data related to Piwi knockdown study portion.
Project description:Many animal species employ a chromosome-based mechanism of sex determination, which has led to coordinate evolution of dosage compensation systems. Dosage compensation not only corrects the imbalance in the number of X-chromosomes between the sexes, but is also hypothesized to correct dosage imbalance within cells due to mono-allelic X expression and bi-allelic autosomal expression, by upregulating X-linked genes (termed M-bM-^@M-^XOhnoM-bM-^@M-^Ys hypothesisM-bM-^@M-^Y). To identify molecular mechanisms of X upregulation in mammals we established genome-wide profiles for the initiation and elongation forms of RNA polymerase II (PolII), PolII-S5p (phosphorylated at serine 5) and PolII-S2p (phosphorylated at serine 2), and for histone modifications in mouse cell lines and tissues. We found that in addition to being enriched in PolII-S5p but not in PolII-S2p, X-linked promoters were also enriched in two epigenetic marks, H4K16ac and H2AZ, dependent on expression levels. To address the function of the H4K16 acetyltransferase MOF occupancy profiles were established and knockdowns of MOF and MSL1 were done in mouse ES cells. Our results support a conserved role for the MSL complex to enhance transcription initiation of X-linked genes. Comparison of the profiles of RNA PolII, the H4K16ac acetyltransferase MOF and histone active marks on the X versus autosomes in mouse