ABSTRACT: H4K12 acetylation mapped by chromatin Chromatin immunoprecipitation (ChIP) in human sperm. ChIP mapping of chromatin modification. Three biological replicates hybridized to each of two chips.
During the haploid phase of mammalian spermatogenesis, nucleosomal chromatin is ultimately repackaged by small, highly basic protamines to generate an extremely compact, toroidal chromatin architecture that is critical to normal spermatozoal function. In common with several species, however, the human spermatozoon retains a small proportion of its chromatin packaged in nucleosomes. As nucleosomal chromatin in spermatozoa is structurally more open than protamine-packaged chromatin, we considered ...[more]
Project description:Forkhead box A2 (FOXA2) is a critical regulator of endometrial gland development in mice. In the adult mouse uterus, FOXA2 is expressed solely in the GE cells of the endometrium. Conditional deletion of Foxa2 after birth in the uterus, using the progesterone receptor Cre mouse (PgrCre), impeded gland development, thereby rendering the adult mouse infertile due to defects in blastocyst implantation stemming from a lack of endometrial glands and their secretions. As a first step to begin understanding the FOXA2 function in the endometrial glands of the uterus, genome-wide investigation of in vivo FOXA2 and RNA polymerase II (POL2) binding target regions in the neonatal and adult uterus was determined by chromatin immunoprecipitation followed by massively parallel sequencing (ChIP-Seq). In order to determine the transcriptional regulatory networks mediating FOXA2 regulation of endometrial gland development and function, chromatin immunoprecipitation and massively parallel sequencing (ChIP-Seq) was used to create a genome-wide profile of in vivo FOXA2-binding sites in the developing (PD 12) and adult (DOPP 2.5 and 3.5) mouse uterus.
Project description:Regulation of Megakaryocytic differentiation in Cell Line Models by Dynamic Combinatorial Interactions of RUNX1 with Its Cooperating Partners Examination of RUNX1 binding in K562 cells, before and following TPA induction and CMK cells. Examination of GATA1 and FOS binding and H3K4me1 and H3K27me3 modification levels following TPA induction in K562 cells.
Project description:R-loops are transcription by-products that may constitute a threat to genome integrity. In addition to specific enzymes to remove them, eukaryotes rely on a number of mRNP biogenesis factors such as the THO complex, to prevent co-transcriptional R-loop formation. We show in Saccharomyces cerevisiae that R-loops are tightly and specifically linked with histone H3-Ser10 phosphorylation (H3S10P), a mark of chromatin condensation. Importantly, ChIP-chip analyses reveal a clear H3S10P accumulation at the pericentromeric chromatin during the G1-phase of the cell cycle only in R loop-accumulating yeast strains but not in those non-accumulating R-loops, and a significantly higher accumulation during S-phase. Such a difference can also be detected in a number of genes along the genome. ChIP-chip studies were perfomed with antibodies against Histone H3 and the phosphorylated Histone H3 at Serine10 in the yeast S. cerevisiae.
Project description:R-loops are transcription by-products that may constitute a threat to genome integrity.We previously show in Saccharomyces cerevisiae that R-loops are tightly and specifically linked with histone H3-Ser10 phosphorylation (H3S10P), a mark of chromatin condensation. Here we analyse the hpr1-101 mutant. A point mutation that impairs transcription and mRNP biogenesis without increasing recombination. Importantly, ChIP-chip analyses reveal a clear H3S10P does not accumulate at the pericentromeric chromatin during the G1-phase of the cell cycle but during S-phase in non-accumulating R-loop hpr1-101. ChIP-chip studies were perfomed with antibodies against Histone H3 and the phosphorylated Histone H3 at Serine10 in the yeast S. cerevisiae.
Project description:The Fil1 transcription factor regulates the response to amino acid starvation. Here we analyse the effects of overexpressing the fil1 coding sequence and of its deregulated expression. The following strains were used: 1] Strains expressing the fil1 gene under the control of the thiamine-repressible nmt1 promoter (nmt-fil1) 2] Strains in which 6 uORFs (upstream Open Reading Frames) in the fil1 5'-UTR have been inactivated, leading to deregulated translation of the fil1 mRNA (6uORF_mutant)
Project description:Purpose: Description of a spike-adjusting-method (SAM) to normalize ChIP-seq data . Methods: We performed ChIP-seq of POLR3D and POLR2B with mouse liver supplemented with 2.5% of human DNA. Human DNA will be used as an internal control for ChIP-seq quantification. Results: We show that using the SAM for ChIP-seq quantification improve similarity of POLR3D and POLR2B ChIP-seq replicates samples and improve difference between samples originate from different conditions. Conclusions: The SAM improves comparison of ChIP-seq samples, either by increasing similarity between replicates or by emphasise differences between conditions. Chromatin Immuno-precipitations were performed with antibodies directed against POLR3D (Pol III) and POLR2B (Pol II) using mouse liver material supplemented with human DNA. Immuno-precipitated DNA was next sequenced using Illumina HiSeq. Three different concentrations of human spiked DNA were tested for the Pol III ChIP (2.5%, 5% and 10%). We also sequenced the corresponding inputs (crosslinked DNA from mouse liver). Two concentrations of human spiked DNA (5% and 10%) were tested for the Pol 2 ChIP. We also sequenced the corresponding inputs (crosslinked DNA from mouse liver).