Application of recombinant PHD domain instead of anti-H3K4me3 antibody
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Illumina HiSeq 2500
ORGANISM(S): Homo sapiens
SUBMITTER: Goran Kungulovski
PROVIDER: E-MTAB-4103 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:Application of recombinant PHD domain instead of anti-H3K4me3 antibody
| PRJEB11987 | ENA
Project description:BACKGROUND:Histone posttranslational modifications (PTMs) represent a focal point of chromatin regulation. The genome-wide and locus-specific distribution and the presence of distinct histone PTMs is most commonly examined with the application of histone PTM-specific antibodies. In spite of their central role in chromatin research, polyclonal antibodies suffer from disadvantages like batch-to-batch variability and insufficient documentation of their quality and specificity. RESULTS:To mitigate some of the pitfalls of using polyclonal antibodies against H3K4me3, we successfully validated the application of a recombinant TAF3 PHD domain as anti-H3K4me3 affinity reagent in peptide array, western blot and ChIP-like experiments coupled with qPCR and deep sequencing. CONCLUSIONS:The successful addition of the TAF3 PHD domain to the growing catalog of recombinant affinity reagents for histone PTMs could help to improve the reproducibility, interpretation and cross-laboratory validation of chromatin data.
| S-EPMC4802638 | biostudies-literature
Project description:Histone modifications regulate chromatin-dependent processes, yet the mechanisms by which they contribute to specific outcomes remain unclear. H3K4me3 is a prominent histone mark that is associated with active genes and promotes transcription through interactions with effector proteins that include initiation factor TFIID. We demonstrate that H3K4me3-TAF3 interactions direct global TFIID recruitment to active genes, some of which are p53 targets. Further analyses show that (i) H3K4me3 enhances p53-dependent transcription by stimulating preinitiation complex (PIC) formation; (ii) H3K4me3, through TAF3 interactions, can act either independently or cooperatively with the TATA box to direct PIC formation and transcription; and (iii) H3K4me3-TAF3/TFIID interactions regulate gene-selective functions of p53 in response to genotoxic stress. Our findings indicate a mechanism by which H3K4me3 directs PIC assembly for the rapid induction of specific p53 target genes Examination of genome wide binding sites of TAF3 full length protein vs TAF3 PHD domain alone (Full vs PHD), with or without M880A mutation (WT vs mut) in mouse MEF cells using HITseq method (PNAS 2010, 107:3135-3140, PMID: 20133638)
2013-03-02 | E-GEOD-43540 | biostudies-arrayexpress
Project description:The aim of the experiment was to determine if changes in nuclear phosphoinositides induced by depletion of PIP4K2B impacts on gene expression through a specific phosphoinositide interaction site on TAF3. TAF3 is a core promoter complex protein that contains a PHD finger that interacts with H3K4me3. We determined that The TAF3 PHD finger also interacted with phosphoinositides and generated mutants of TAF3 that were unable to interact with phosphoinositides but still were capable of interacting with H3K4me3. PIP4K2B depletion enhances C2C12 myoblast differentiation. We depleted the endogenous TAF3 from C2C12 myoblast cells and rescued the cells with either a wild type TAF3 or a mutant unable to interact with PI (KK-TAF3). These cells were then maintained as controls or depeleted of PIP4K2B. The cells were then differentiated for two days or were treated wtih a etoposide. We aimed to identify genes that were reguated by PIP4K2B that required an intact phosphoinositide binding site.
2015-02-28 | GSE66353 | GEO
Project description:The aim of the experiment was to determine if changes in nuclear phosphoinositides induced by depletion of PIP4K2B impacts on gene expression through a specific phosphoinositide interaction site on TAF3. TAF3 is a core promoter complex protein that contains a PHD finger that interacts with H3K4me3. We determined that The TAF3 PHD finger also interacted with phosphoinositides and generated mutants of TAF3 that were unable to interact with phosphoinositides but still were capable of interacting with H3K4me3. PIP4K2B depletion enhances C2C12 myoblast differentiation. We depleted the endogenous TAF3 from C2C12 myoblast cells and rescued the cells with either a wild type TAF3 or a mutant unable to interact with PI (KK-TAF3). These cells were then maintained as controls or depeleted of PIP4K2B. The cells were then differentiated for two days or were treated wtih a etoposide. We aimed to identify genes that were reguated by PIP4K2B that required an intact phosphoinositide binding site. Triplicate biological samples were analysed for each point after differentiation for three days or etoposide treatment for seven hours.
2015-02-28 | E-GEOD-66353 | biostudies-arrayexpress
Project description:Histone modifications regulate chromatin-dependent processes, yet the mechanisms by which they contribute to specific outcomes remain unclear. H3K4me3 is a prominent histone mark that is associated with active genes and promotes transcription through interactions with effector proteins that include initiation factor TFIID. We demonstrate that H3K4me3-TAF3 interactions direct global TFIID recruitment to active genes, some of which are p53 targets. Further analyses show that (i) H3K4me3 enhances p53-dependent transcription by stimulating preinitiation complex (PIC) formation; (ii) H3K4me3, through TAF3 interactions, can act either independently or cooperatively with the TATA box to direct PIC formation and transcription; and (iii) H3K4me3-TAF3/TFIID interactions regulate gene-selective functions of p53 in response to genotoxic stress. Our findings indicate a mechanism by which H3K4me3 directs PIC assembly for the rapid induction of specific p53 target genes Examination of TAF3, RNAPII, and H3K4me3 distribution in HCT116 cells.
2013-03-02 | E-GEOD-43539 | biostudies-arrayexpress
Project description:Histone modifications regulate chromatin-dependent processes, yet the mechanisms by which they contribute to specific outcomes remain unclear. H3K4me3 is a prominent histone mark that is associated with active genes and promotes transcription through interactions with effector proteins that include initiation factor TFIID. We demonstrate that H3K4me3-TAF3 interactions direct global TFIID recruitment to active genes, some of which are p53 targets. Further analyses show that (i) H3K4me3 enhances p53-dependent transcription by stimulating preinitiation complex (PIC) formation; (ii) H3K4me3, through TAF3 interactions, can act either independently or cooperatively with the TATA box to direct PIC formation and transcription; and (iii) H3K4me3-TAF3/TFIID interactions regulate gene-selective functions of p53 in response to genotoxic stress. Our findings indicate a mechanism by which H3K4me3 directs PIC assembly for the rapid induction of specific p53 target genes. Total RNAs from control or TAF3 knockdown cells before and after doxorubicin treatment were subjected to Illumina microarray analyses.
2013-03-02 | E-GEOD-43541 | biostudies-arrayexpress
Project description:Histone modifications regulate chromatin-dependent processes, yet the mechanisms by which they contribute to specific outcomes remain unclear. H3K4me3 is a prominent histone mark that is associated with active genes and promotes transcription through interactions with effector proteins that include initiation factor TFIID. We demonstrate that H3K4me3-TAF3 interactions direct global TFIID recruitment to active genes, some of which are p53 targets. Further analyses show that (i) H3K4me3 enhances p53-dependent transcription by stimulating preinitiation complex (PIC) formation; (ii) H3K4me3, through TAF3 interactions, can act either independently or cooperatively with the TATA box to direct PIC formation and transcription; and (iii) H3K4me3-TAF3/TFIID interactions regulate gene-selective functions of p53 in response to genotoxic stress. Our findings indicate a mechanism by which H3K4me3 directs PIC assembly for the rapid induction of specific p53 target genes.
2013-03-02 | GSE43541 | GEO
Project description:Histone modifications regulate chromatin-dependent processes, yet the mechanisms by which they contribute to specific outcomes remain unclear. H3K4me3 is a prominent histone mark that is associated with active genes and promotes transcription through interactions with effector proteins that include initiation factor TFIID. We demonstrate that H3K4me3-TAF3 interactions direct global TFIID recruitment to active genes, some of which are p53 targets. Further analyses show that (i) H3K4me3 enhances p53-dependent transcription by stimulating preinitiation complex (PIC) formation; (ii) H3K4me3, through TAF3 interactions, can act either independently or cooperatively with the TATA box to direct PIC formation and transcription; and (iii) H3K4me3-TAF3/TFIID interactions regulate gene-selective functions of p53 in response to genotoxic stress. Our findings indicate a mechanism by which H3K4me3 directs PIC assembly for the rapid induction of specific p53 target genes
2013-03-02 | GSE43540 | GEO
Project description:Histone modifications regulate chromatin-dependent processes, yet the mechanisms by which they contribute to specific outcomes remain unclear. H3K4me3 is a prominent histone mark that is associated with active genes and promotes transcription through interactions with effector proteins that include initiation factor TFIID. We demonstrate that H3K4me3-TAF3 interactions direct global TFIID recruitment to active genes, some of which are p53 targets. Further analyses show that (i) H3K4me3 enhances p53-dependent transcription by stimulating preinitiation complex (PIC) formation; (ii) H3K4me3, through TAF3 interactions, can act either independently or cooperatively with the TATA box to direct PIC formation and transcription; and (iii) H3K4me3-TAF3/TFIID interactions regulate gene-selective functions of p53 in response to genotoxic stress. Our findings indicate a mechanism by which H3K4me3 directs PIC assembly for the rapid induction of specific p53 target genes
2013-03-02 | GSE43539 | GEO