Project description:Setd2 is the specific methyltransferase of H3K36me3. To understand the global effect of H3K36me3 on m6A modification, we used mouse embryonic stem cells (mESCs) model with doxycycline (Dox)-induced Setd2 knockdown, and performed m6A-IP followed by sequencing in mESCs with or without Dox treatment. We found that depletion of H3K36me3 by Setd2 silencing globally reduced m6A in mouse transcriptome.
Project description:SETD2 is the specific methyltransferase of H3K36me3, while METTL3, METTL14 and WTAP are the components of m6A methyltransferase complex. To understand the global effect of H3K36me3 on m6A modification, we compared the m6A profiling in SETD2 and METTL3, METTL14 or WTAP knockdown HepG2 cells, and found depletion of H3K36me3 by SETD2 silencing globally reduced m6A in human transcriptome. What’s more, most of the SETD2-dependent hypomethylation sites also responded to knockdown of METTL3, METTL14, or WTAP.
Project description:To understand the global effect of H3K36me3 on m6A modification, we compared the m6A profiling in SETD2 knockdown and control HepG2 cells by miCLIP-seq, and found the depletion of H3K36me3 by SETD2 silencing globally reduced m6A in the human transcriptome.
Project description:H3K36me3 has been reported to associate with active gene expression, and SETD2 is the mainly methyltransferase for H3K36me3. We identified SPOP which is a CUL3 family protein as a E3 ligase for SETD2. Genome wide analysis by using ChIPSeq and RNASeq demonstrate that SPOP specificly eliminate H3K36me3 modification at target genes and resulted in alternative splicing of those target genes.
Project description:Spermatogenesis is precisely cotrolled by complex gene expression programs and involves epigenetic reprogramming including histone modification and DNA methylation. Setd2 catalyzes the trimethylation of histone H3 Lys36 (H3K36me3) and plays key roles in embryonic stem cell differentiation and somatic cell development; however, its role in male germ cell development remains elusive. Here we demonstrate an essential role of Setd2 for spermiogenesis. We show that targeted knockout of Setd2 in germ cells causes aberrant spermiogenesis with acrosomal malformation before step 8 round spermatid stage, resulting in complete male infertile. Furthermore, we show a complete loss of H3K36me3 and a significant altered gene expression profile, including Acrbp1 and protamines, caused by Setd2 deficiency. Our findings reveal a previously underappreciated role of Setd2-dependent H3K36me3 for spermiogenesis and improved the understanding of epigenetic disorders underlying male infertility.
Project description:We report the application of H3K36me3 ChIP sequencing in SETD2 genotyped samples Examination of H3K36me3 in SETD2 wild-type, mutant renal cell carcinoma and SETD2 isogenic cell lines
Project description:Several lines of recent evidence support a role for chromatin in splicing regulation. Here we show that splicing can also contribute to histone modification, which implies a bidirectional communication between epigenetics and RNA processing. Genome-wide analysis of histone methylation in human cell lines and mouse primary T cells reveals that intron-containing genes are preferentially marked with H3K36me3 relative to intronless genes. In intron-containing genes, H3K36me3 marking is proportional to transcriptional activity, whereas in intronless genes H3K36me3 is always detected at much lower levels. Furthermore, splicing inhibition impairs recruitment of H3K36 methyltransferase HYPB/Setd2 and reduces H3K36me3, whereas splicing activation has the opposite effect. Moreover, the increase of H3K36me3 correlates with the length of the first intron, consistent with the view that splicing enhances H3 methylation. We propose that splicing is mechanistically coupled to recruitment of HYPB/Setd2 to elongating RNA Polymerase II. This experiment proposes to profile genome-wide binding profiles by ChIP-seq (Illumina, 36 bp tags) of RNA polymerase II (one biological replicate), the histone modification H3K36me3 (2 replicates) and a reference control input sample (genomic DNA after reverse cross-link, one replicate) in a human H1299 lung carcinoma cell line *** Raw data not provided for Samples GSM766322-GSM766324.
Project description:SETD2 is the sole chromatin modifier responsible for H3K36me3, a histone mark linked to splicing, transcription initiation and DNA damage response. Homozygous disruption of SETD2 yields a tumor suppressor effect in various cancers. However, SETD2 mutation is virtually always heterozygous in diffuse large B-cell lymphomas (DLBCL). Here we show that heterozygous SETD2 deficiency results in germinal center (GC) hyperplasia and accelerated lymphomagenesis. SETD2 haploinsufficient GC B-cells exhibit increased competitive fitness and reduced DNA damage checkpoint activity, resulting in decreased apoptosis. SETD2 haploinsufficient GCB and lymphoma cells featured increased off- and on-target AICDA induced somatic hypermutation (SHM), complex structural variants such as rygma, and increased translocations including those activating MYC. DNA damage was selectively increased on the non-template strand and H3K36me3 loss was associated with greater RNA Pol II processivity and mutational burden, suggesting that SETD2-mediated H3K36me3 is required for proper sensing of cytosine deamination during transcription. Hence, SETD2 haploinsufficiency delineates a novel GC B-cell context specific oncogenic pathway involving defective epigenetic surveillance of AICDA mediated somatic hypermutation induced off target effects on transcribed genes.
Project description:Genome-wide analysis of histone modification (H2AZ, H3K27ac, H3K27me3, H3K36me3, H3K4me1, H3K4me2, H3K4me3 and H3K9me3), protein-DNA binding (TAF1, P300, Pou5f1 and Nanog), cytosine methylation and transcriptome data in mouse and human ES cells and pig iPS cells We generated histone modification data (H2AZ, H3K27ac, H3K27me3, H3K36me3, H3K4me1, H3K4me2, H3K4me3 and H3K9me3) and protein-DNA binding data (TAF1, P300, Pou5f1 and Nanog) using Chromatin Immunoprecipitation followed by short sequencing (ChIP-seq), cytosine methylation data using methylated DNA immunoprecipitation followed by sequencing (MeDIP-seq) and DNA digestion by methyl-sensitive restriction enzymes followed by sequencing (MRE-seq), transcriptome data with RNA short sequencing (RNA-seq) in human embryonic stem cells, mouse embryonic stem cells, pig induced pluripotent stem cells and mouse embryonic stem cells under activin-A-induced-differentiation.
