Project description:The KDM4/JMJD2 are H3K9- and H3K36- specific demethylases, which are considered promising therapeutic targets for the treatment of acute myeloid leukemia (AML) harboring MLL-translocations. Here, we investigate the long-term effects of depleting KDM4 activity on normal hematopoiesis to probe potential side effects of continuous inhibition of these enzymes. Utilizing conditional Kdm4a/Kdm4b/Kdm4c triple-knockout mice we show that KDM4 activity is required for hematopoietic stem cell (HSC) maintenance in vivo. The knockout of the KDM4 demethylases leads to accumulation of H3K9me3 on transcription start sites and the corresponding downregulation of expression of several genes in hematopoietic stem cells. We show that two of these genes, Taf1b and Nom1, are essential for the maintenance of hematopoietic cells. Taken together, our results show that the KDM4 demethylases are required for the expression of genes essential for the long-term maintenance of normal hematopoiesis.
Project description:We report the application of low input high-throughput profiling of histone modifications in mouse oocytes. Using antibodies against H3K4me3 (100 oocytes/replicate; 2 biological replicates per genotype) and H3K9me3 ( around 300 oocytes/replicate; 2 biological replicates per genotype), we find that they are mutually exclusive in oocytes and this property is lost in oocytes lacking Kdm4a. H3K9me3 is spread into regions of H3K4me3 postive open chromatin which is surprisingly, well preserved leading to a potential bivalency of scale. Altogether, KDM4A is important to keep H3K4me3 marked open chromatin clear of H3K9me3 spreading in oocytes.
Project description:ETV2/ER71, an ETS transcription factor, is critical for hematopoiesis and vascular development. However, knowledge on the molecular mechanisms behind ETV2-mediated gene transcription is limited. Here, we show that ETV2 together with KDM4A, an H3K9 demethylase, regulates hematopoietic and endothelial genes. Etv2-/- mouse embryonic stem cells (mESCs), which fail to generate hematopoietic and endothelial cells, showed enhanced levels of H3K9me3 on hematopoietic and endothelial genes. ETV2 interacts with KDM4A and the ETV2-mediated transcriptional activation of hematopoietic and endothelial genes is dependent on KDM4A histone demethylase activity. ETV2 and KDM4A co-occupy the transcription regulatory regions of genes whose expression is directly regulated by ETV2. Mice lacking Kdm4a and Etv2 in endothelial cells (Cdh5Cre;Kdm4af/f;Etv2f/f) displayed a more severe defect in perfusion recovery and neovascularization compared with Cdh5Cre;Kdm4af/f, Cdh5Cre;Etv2f/f mice and controls. Collectively, we demonstrated that ETV2 interacts with KDM4A and that this interaction is critical for FLK1+ cell generation, differentiation into the downstream lineages, and vascular regeneration.
Project description:We report the application of low input high-throughput profiling of histone modifications in mouse oocytes and 2-cell embryos. We used antibodies against H3K36me3 (190 wildtype and 93 knockout MIIcoocytes pooled as one replicate) and H3K9me3 ( around 118 control and MZ embryos / replicate; 1 biological replicates per genotype). In the case of H3K36me3, we find that its establishment and maintence is not affected in oocytes lacking Kdm4a. In the case of H3K9me3 in 2-cell embryos, we find that the histone mark aberrantly gained in Kdm4a deficient oocytes is also maintained in the MZ mutant 2-cell embryo. This shows that Kdm4a is important to keep open chromatin clear of H3K9me3 spreading in oocytes in order to transfer a healthy epigenome amenable for proper genome activation post fertilization.
Project description:Purpose: ChipSeq allows us to identify binding sites of target proteins of interest in the genome, helping create a global profile for the same. Kdm4a is a H3K9me3 histone demethylase that acts to prevent spurious accumulation of this repressive mark at H3K4me3 positive transcription start sites. To complement transcriptome data from Kdm4a mice, we performed Chip-Seq on 2.5dpc mouse uterus of 8-10 week old littermate control and Kdm4a knockout mice for Kdm4a, H3K9me3 and H3K4me3. This would help us check which of the target genes with changed gene expression was due to the direct change in histone lanscape in absence of Kdm4a.
Project description:To explore the underlying mechanism of KDM4A mediated H3K9me3 modification in CRC. We performed Chromatin immunoprecipitation sequencing(CHIP-seq) between control CRC cells and CRC cells with KDM4A overexpression to identify KDM4A regulating signaling pathway.
Project description:We previously identified an essential role for the H3K9me3 histone demethylase, KDM4A, in maintaining AML cell survival, with genetic depletion by shRNA of KDM4A inducing leukaemic cell death with little effect on normal haematopoiesis. We hypothesised KDM4A inhibition may represent a novel and effective strategy to treat AML. To address this, we provided proof-of-principle for the use of KDM4 inhibitors (KDM4Ai) and fully characterised their functional potential in AML cells and identified the epigenetic mechanisms underpinning the essential role of KDM4A activity in THP-1 cells using both ChIP-seq and RNA-seq, delineating and functionally validating the epigenomic network regulated by KDM4A, showing that selective loss of KDM4A is sufficient to induce apoptosis in a broad spectrum of human AML cells. This detrimental phenotype results from a global accumulation of H3K9me3 and H3K27me3 at KDM4A targeted genomic loci thereby causing down-regulation of a KDM4A-PAF1 controlled transcriptional program essential for leukemogenesis, distinct from that of KDM4C.
Project description:The importance of germline-inherited posttranslational histone modification in priming early mammalian development is just emerging1-4. Histone H3 lysine 9 (H3K9) trimethylation is associated with heterochromatin and gene repression during cell-fate change5, while histone H3 lysine 4 (H3K4) trimethylation marks active gene promoters6. Mature oocytes are transcriptionally quiescent and possess remarkably broad domains of H3K4me3 (bdH3K4me3)1, 2. It remains unknown, which factors contribute to the maintenance of the bdH3K4me3 landscape. Lysine-specific demethylase 4A (KDM4A) demethylates H3K9me3 at promoters marked by H3K4me3 in actively transcribing somatic cells7. Here, we report that KDM4A-mediated H3K9me3 demethylation at bdH3K4me3 in oocytes is crucial for normal preimplantation development and zygotic genome activation (ZGA) after fertilization. Loss of KDM4A in oocytes causes extensive and aberrant H3K9me3 spreading at bdH3K4me3, resulting in insufficient transcriptional activation ZGA genes, endogenous retroviral elements and long terminal repeat -initiated chimeric transcripts in 2-cell embryos. The catalytic activity of KDM4A is essential for normal epigenetic reprogramming and preimplantation development. Hence, KDM4A plays a crucial role in preserving maternal epigenome integrity required for proper ZGA and transfer of developmental control to the embryo.
Project description:Isocitrate dehydrogenases 1 and 2 are mutated in more than 70% of low-grade glioma. This promotes the production of R-2-hydroxyglutarate (R-2HG) instead of α-ketoglutarate (αKG) and R-2HG will inhibit α-ketoglutarate enzymes like the lysine demethylase KDM4A. We hypothesized that KDM4A inhibition through R-2HG contribute to gliomagenesis. In our study, we showed that KDM4A inhibition leads to telomere defects. By ChIP-seq experiments, we demonstrated that KDM4A localized on telomeric repeats and that its depletion increased the level of H3K9me3 on telomeric chromatin.