DAXX genome binding/occupancy profiling by high throughput ChIP sequencing
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ABSTRACT:
PROVIDER: PRJNA788463 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR17217593.fastq.gz | Fastqsanger.gz | |||
| SRR17217594.fastq.gz | Fastqsanger.gz | |||
| SRR23249045.fastq.gz | Fastqsanger.gz |
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Project description:Purpose: The goal of this study is to compare DAXX genome binding/occupancy by high throughput ChIP sequencing in control, wild type DAXX overexpressd cells (WT DAXX OE), and wild type DAXX SUMO interacting motif mutant overexpressd cells (DSM OE). Methods: The panel of MDA_MB-231 derived cell lines (control, WT DAXX OE, DSM OE) were cultured in complete DMEM. At about 90% confluency, the cells were crosslinked by adding 37 % formaldehyde to the final concentration of 1% for 10 min at room temperature. Crosslinking was stopped by adding glycine to the final concentration of 125 mM. Cells were lifted, washed with cold PBS, and pelleted by centrifugation. The cells were resuspended in a swelling buffer in the presence of the protease inhibitor cocktail (Sigma) and then pelleted and resuspended in the SDS lysis buffer. The lysates were transferred to a Covaris microTUBE and sonicated with an E220 Covaris Ultrasonicator. Chromatin fragmentation (~500 bps) was verified through agarose gel electrophoresis. The fragmented chromatins were diluted and incubated with a control IgG and the DAXX mAb (5G11) along with protein A/G magnetic beads. The beads were washed sequentially with a low salt buffer, high salt buffer, LiCl buffer, and TE buffer (twice). The immunoprecipitated chromatins were eluted at 65 °C for 15 min, and the eluted chromatins were subjected to proteinase K digestion at 65 °C for 3 h. The DNAs were recovered through a Qiagen mini-prep column. The immunoprecipitated DNAs were used for qPCR and library construction and high throughput sequencing using an Illumina Hi-Seq 2500 sequencer. Results: We surveyed genome-wide occupancy of DAXX using ChIP-seq technology. Overexpression of WT DAXX increased DAXX?s chromatin association specifically with lipogenic pathway genes where DSM OE reverse. Conclusions: Our study represents the first detailed analysis of DAXX occupancy in lipoegenic gene transciption.
2021-12-20 | GSE190783 | GEO
Project description:DAXX, originally discovered as a context-dependent regulator of cell death or survival. Emerging evidence suggests that DAXX has an oncogenic role in diverse cancer types. However, the molecular mechanisms underlying DAXX's oncogenic function remain to be defined. We used high throughput Illumina sequencing to analyze genes that were differentially expressed upon DAXX knockdown or DAXX OE or DSM OE.
2021-12-24 | GSE192420 | GEO
Project description:DAXX and ATRX are tumor suppressor proteins that form a complex with histone H3.3 chaperone and are frequently mutated in cancers with the alternative lengthening of telomeres (ALT), such as pediatric glioblastoma. Rapid loss of function of either DAXX or ATRX are not by themselves sufficient to induce the ALT phenotype. However, cells lacking DAXX or ATRX can be readily selected for ALT-like features. Here, we show that a key feature of ALT selected DAXX and ATRX null glioblastoma cells is the attenuation of p53 function. RNA-seq analysis of DAXX or ATRX null U87 glioblastoma cells with ALT-like features revealed that p53 pathway is among perturbed. ALT-selected DAXX and ATRX-null cells had aberrant response to DNA damaging agent etoposide. Both DAXX and ATRX-null ALT cells showed a loss of p53 binding at a subset of response elements. Complementation of DAXX null cells with wt DAXX rescued p53 binding and transcription, while the tumor associated mutation L130R, that disrupts ATRX binding, was incapable of rescuing p53 chromatin binding. We show that histone H3.3 binding is reduced in DAXX-null cells especially at subtelomeric p53 binding sites and telomere repeats. These findings indicate that DAXX and ATRX function to enable p53 chromatin binding through modulation of histone H3.3 binding, especially at sub-telomeric sites.
2022-07-19 | GSE186442 | GEO
Project description:We report the genome-wide occupancy of exogenous DAXX (Myc-tagged), exogenous full length CBX2.1 (tagged with GAL4 DNA binding domain) and exogenous splice variant CBX2.2 (tagged with GAL4 DNA binding domain) in transiently transfected HEK293 cells. We observe that CBX2.1 but not CBX2.2 can efficiently target DAXX to chromatin.
2020-05-12 | GSE107348 | GEO
Project description:DAXX and ATRX are tumor suppressor proteins that form a complex with histone H3.3 chaperone and are frequently mutated in cancers with the alternative lengthening of telomeres (ALT), such as pediatric glioblastoma. Rapid loss of function of either DAXX or ATRX are not by themselves sufficient to induce the ALT phenotype. However, cells lacking DAXX or ATRX can be readily selected for ALT-like features. Here, we show that a key feature of ALT selected DAXX and ATRX null glioblastoma cells is the attenuation of p53 function. RNA-seq analysis of DAXX or ATRX null U87 glioblastoma cells with ALT-like features revealed that p53 pathway is among perturbed. ALT-selected DAXX and ATRX-null cells had aberrant response to DNA damaging agent etoposide. Both DAXX and ATRX-null ALT cells showed a loss of p53 binding at a subset of response elements. Complementation of DAXX null cells with wt DAXX rescued p53 binding and transcription, while the tumor associated mutation L130R, that disrupts ATRX binding, was incapable of rescuing p53 chromatin binding. We show that histone H3.3 binding is reduced in DAXX-null cells especially at subtelomeric p53 binding sites and telomere repeats. These findings indicate that DAXX and ATRX function to enable p53 chromatin binding through modulation of histone H3.3 binding, especially at sub-telomeric sites.
2022-07-19 | GSE207046 | GEO
Project description:DAXX and ATRX are tumor suppressor proteins that form a complex with histone H3.3 chaperone and are frequently mutated in cancers with the alternative lengthening of telomeres (ALT), such as pediatric glioblastoma. Rapid loss of function of either DAXX or ATRX are not by themselves sufficient to induce the ALT phenotype. However, cells lacking DAXX or ATRX can be readily selected for ALT-like features. Here, we show that a key feature of ALT selected DAXX and ATRX null glioblastoma cells is the attenuation of p53 function. RNA-seq analysis of DAXX or ATRX null U87 glioblastoma cells with ALT-like features revealed that p53 pathway is among perturbed. ALT-selected DAXX and ATRX-null cells had aberrant response to DNA damaging agent etoposide. Both DAXX and ATRX-null ALT cells showed a loss of p53 binding at a subset of response elements. Complementation of DAXX null cells with wt DAXX rescued p53 binding and transcription, while the tumor associated mutation L130R, that disrupts ATRX binding, was incapable of rescuing p53 chromatin binding. We show that histone H3.3 binding is reduced in DAXX-null cells especially at subtelomeric p53 binding sites and telomere repeats. These findings indicate that DAXX and ATRX function to enable p53 chromatin binding through modulation of histone H3.3 binding, especially at sub-telomeric sites.
2022-07-19 | GSE186440 | GEO
Project description:This study was aimed at understanding the genome-wide binding and regulatory role of the DAXX transcriptional repressor, recently implicated in PCa. ChIP-Seq analysis of genome-wide distribution of DAXX in PC3 cells revealed over 59,000 DAXX binding sites, found at regulatory enhancers and promoters. ChIP-Seq analysis of DNA methyltransferase 1 (DNMT1), which is a key epigenetic partner for DAXX repression, revealed that DNMT1 binding was restricted to a small number of DAXX sites. DNMT1 and DAXX bound close to transcriptional activator motifs. DNMT1 sites were found to be dependent on DAXX for recruitment by analyzing DNMT1 ChIP-Seq following DAXX knockdown (K/D), corroborating previous findings that DAXX recruits DNMT1 to repress its target genes. Massively parallel RNA sequencing (RNA-Seq) was used to compare the transcriptomes of WT and DAXX K/D PC3 cells. Genes induced by DAXX K/D included those involved in autophagy, and DAXX ChIP-Seq peaks were found close to the transcription start sites (TSS) of autophagy genes, implying they are more likely to be regulated by DAXX. To determine DAXX binding sites in the prostate cancer (PCa) genome, the PC3 cell line was used. A stable DAXX shRNA knockdown (K/D) PC3 cell line and a control shRNA counterpart, were compared in a ChIP-Seq study.
2015-06-24 | E-GEOD-68647 | biostudies-arrayexpress
Project description:High throughput sequencing for YTHDC1 binding RNAs [ChIP-seq]
| PRJNA610621 | ENA
Project description:Our previous studies have implicated CHIP as a co-chaperone/ubiquitin ligase, whose activities yield protection against stress-induced apoptotic events. In this report, we demonstrate a stress-dependent interaction between CHIP (carboxyl terminus of Hsp70-interacting protein) and Daxx, death domain-associated protein. This interaction interferes with the stress-dependent association of HIPK2 with Daxx, blocking phosphorylation of serine 46 in p53 and inhibiting the p53-dependent apoptotic program. Microarray analysis confirmed suppression of the p53-dependent transcriptional portrait in CHIP (+/+) but not in CHIP (-/-) heat shocked MEFs. The interaction between CHIP and Daxx results in ubiquitination of Daxx which is then partitioned to an insoluble compartment of the cell. In vitro ubiquitination of Daxx by CHIP revealed that Ub chain formation utilizes non canonical lysine linkages associated with resistance to proteasomal degradation. CHIP's ubiquitination of Daxx utilizes lysines 630 and 631 and competes with the cell's sumoylation machinery at these residues. These studies implicate CHIP as a stress-dependent regulator of Daxx that counters Daxx's pro-apoptotic influence in the cell. By abrogating p53-dependent apoptotic pathways and by ubiquitination competitive with Daxx sumoylation, CHIP integrates the cell's proteotoxic stress response with cell cycle pathways that influence cell survival. Keywords: p53, apoptosis, cell stress, ubiquitination We utilized a âsample x referenceâ experimental design strategy in which RNA extracted from mouse embryonic fibroblasts was hybridized to the microarray slide in the presence of labeled Universal Mouse Reference RNA (UMRR, Stratagene, LaJolla, CA). A total of 24 RNA samples were used in this analysis. Briefly, five hundred nanograms of total RNA were used for gene expression profiling following reverse transcription and T-7 polymerase-mediated amplification/labeling with Cyanine-5 CTP. Labeled subject cRNA was co-hybridized to Agilent G4112F Whole Mouse Genome 4x44K oligonucleotide arrays with equimolar amounts of Cyanine-3 labeled UHRR. Slides were hybridized, washed, and scanned on an Axon 4000b microarray scanner. The data were processed using Feature Extaction software (Agilent, Santa Clara, CA).
2010-04-03 | E-GEOD-14339 | biostudies-arrayexpress
Project description:Using ChIP-seq to profile the genomic binding of DAXX in control and C9HRE ALS/FTD patient B lymphocytes.
2023-02-10 | GSE223401 | GEO