MIR sequences recruit zinc finger protein ZNF768 to expressed genes
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA438474 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR6841440.fastq.gz | Fastqsanger.gz | |||
| SRR6841441_1.fastq.gz | Fastqsanger.gz | |||
| SRR6841441_2.fastq.gz | Fastqsanger.gz | |||
| SRR6841442_1.fastq.gz | Fastqsanger.gz | |||
| SRR6841442_2.fastq.gz | Fastqsanger.gz |
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Project description:Mammalian-wide interspersed repeats (MIRs) are retrotransposed elements of mammalian genomes. Here we report the specific binding of zinc finger protein ZNF768 to the sequence motif GCTGTGTG (N20) CCTCTCTG in the core region of MIRs. ZNF768 binding is preferentially associated with euchromatin and promoter regions of genes. Binding was observed for genes expressed in a cell type-specific manner in human B cell line Raji and osteosarcoma U2OS cells. Mass spectrometric analysis of ZNF768 associated proteins revealed binding of ZNF768 to Elongator subunits 1, 2, and 3. The N-terminus of ZNF768 contains a heptad repeat array structurally related to the C-terminal domain (CTD) of RNA polymerase II. This array evolved in placental animals but not marsupials and monotreme species, displays species-specific length variations, and possibly fulfills CTD related functions in gene regulation. We propose that the evolution of MIRs and ZNF768 has extended the repertoire of gene regulatory mechanisms in mammals and that ZNF768 binding is associated with cell type-specific gene expression.
2018-10-29 | PXD010831 | Pride
Project description:Mammalian-wide interspersed repeats (MIRs) are retrotransposed elements of mammalian genomes. Here, we report the specific binding of zinc finger protein ZNF768 to the sequence motif GCTGTGTG (N20) CCTCTCTG in the core region of MIRs. ZNF768 binding is preferentially associated with euchromatin and promoter regions of genes. Binding was observed for genes expressed in a cell type-specific manner in human B cell line Raji and osteosarcoma U2OS cells. Mass spectrometric analysis revealed binding of ZNF768 to Elongator components Elp1, Elp2 and Elp3 and other nuclear factors. The N-terminus of ZNF768 contains a heptad repeat array structurally related to the C-terminal domain (CTD) of RNA polymerase II. This array evolved in placental animals but not marsupials and monotreme species, displays species-specific length variations, and possibly fulfills CTD related functions in gene regulation. We propose that the evolution of MIRs and ZNF768 has extended the repertoire of gene regulatory mechanisms in mammals and that ZNF768 binding is associated with cell type-specific gene expression.
| S-EPMC6344866 | biostudies-literature
Project description:U2OS osteosarcoma cells were cultured in Dulbecco's modified Eagle's medium (DMEM, Gibco) and Raji B-cells in RPMI 1640 medium (Gibco) supplemented with 10% fetal calf serum (FCS, Bio&Sell), 2 mM L-glutamine (Gibco), 100 U/mL penicillin (Gibco), and 100 µg/mL streptomycin (Gibco) at 37°C at 8% or 5% CO2, respectively. Chromatin immunoprecipitation for ChIP-seq: Cells were crosslinked using a formaldehyde containing solution (10mM NaCl, 0,1mM EDTA pH 8, 0,05mM EGTA pH 8, 5mM HEPES pH 7.8 and 1% formaldehyde) for 10 minutes at 20°C, the reaction was quenched by the addition of glycin to a final concentration of 250uM for 5 minutes. Crosslinked cells were collected and washed twice with PBS before snap freezing in liquid nitrogen and storage at -80°C until subsequent use. Prior to sonication, the crosslinked cells were resuspended in lysis buffer (50mM Hepes pH 7.5, 140mM NaCl, 1mM EDTA pH 8, 10% glycerol, 0.75% NP-40, 0.25% Triton X-100, 1X protease inhibitor cocktail) at 4°C for 20 minutes. Nuclei were collected by centrifugation and washed in a second buffer (200mM NaCl, 1mM EDTA pH 8, 0.5mM EGTA pH 8, 10mM Tris pH 8, 1X protease inhibitor cocktail) for 10 minutes at 4°C then collected by centrifugation and resuspended in the shearing buffer (1mM EDTA pH 8, 0.5mM EGTA pH 8, 10mM Tris pH 8, 100mM NaCl, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine, 1X protease inhibitor cocktail). Sonication was carried out in a Bioruptor Pico ultrasounds water bath (Diagenode B01060001) for 30 cycles of 30 sec ON and 30 sec OFF pulses in 4°C water. Sonicated extracts were centrifuged at high speed in the presence of 0,1% of Triton X-100 and snap frozen in liquid nitrogen then stored at -80°C until subsequent use. Prior to ChIP, the ZNF768 mAb was coupled to protein-G coated magnetic beads (Dynabeads, life technologies) by incubation in 0,5% BSA PBS overnight at 4°C. Pre-coated beads were then washed and incubated with the sonicated chromatin extracts, the ChIP was carried out overnight at 4°C on a rotating wheel. The equivalent of 10 × 107 cells sonicated extract was used for each ZNF ChIP experiment for both cell lines. After incubation, the beads were washed 7 times with Wash buffer (50mM Hepes pH 7.6, 500mM LiCl, 1mM EDTA pH 8, 1% NP-40, 0.7% Na-Deoxycholate, 1X protease inhibitor cocktail) followed by one wash with TE-NaCl buffer (10mM Tris pH 8 and 1mM EDTA pH 8, 50mM NaCl). Immunoprecipitated chromatine was eluted by two sequential incubations with 100µl Elution buffer (50mM Tris pH 8, 10mM EDTA pH 8, 1% SDS) at 65°C for 15 minutes. The two eluates were pooled and incubated at 65°C for 12 hours to reverse-crosslink the chromatin followed by treatment with RNase A (0.2µg/mL) at 37°C for two hours and Proteinase K (0.2µg/mL) at 55°C for two hours. The DNA was isolated by phenol:chloroform: isoamylalcohol (25:24:1 pH 8) extranction followed by Qiaquick PCR Purification (Qiagen, Germany) and quantified with Qubit DS DNA HS Assay (ThermoFisher Scientific, USA). At least 1ng of ChIP DNA was used to prepare sequencing library with Illumina ChIP Sample Library Prep Kit (Illumina, USA) with a few optimizations to the protocol. The ChIP DNA was size selected using Ampure beads (Life technologies) to enrich for fragments < 400Bp prior to end-repair, 3’end adenylation and adapter ligation. Library fragments were then directly amplified by 10 cycles of PCR.
2018-10-29 | GSE111879 | GEO
Project description:To investigate the collective role of zinc finger proteins in adipogenesis, we induced adipogenesis using human mesenchymal stem cells (MSC) and compared the gene expression profiles using human Illumina beads arrays. We performed three independent experiments of time-series analysis (0h, 1h, 12h, 24h, 2d, 4d, 6d, 9d, 12d, 15d after induction of adipogenesis) in order to comprehensively capture the dynamic profiles of transcriptome during differentiation. Statistical filtering based on Analysis of Variance (ANOVA; p-value < 0.001, FDR 5%) and Gene Ontology classification using the Human Gene Nomenclature Database (HGNC) identified 618 out of 678 zinc finger proteins (ZFPs) that were significantly expressed during adipogenesis. Moreover, 34 out of 618 genes were differentially expressed more than 2-fold up or down regulated. The K-means clustering analysis further revealed four dynamic expression patterns: one gene in early-response, seven genes in transient, twenty genes in progressively-induced and four genes in the down-regulated, demonstrating the dynamic involvement of ZFPs in the process of adipocyte differentiation and maturation. Total RNA obteind from 10 different time points (0h, 1h, 12h, 24h, 2d, 4d, 6d, 9d, 12d, 15d) during adipogenesis from human Mesenchymal stem cell (MSC) to Adipocyte (ADP). Adipogensis was induced by the different cocktail of chemical compunds containing human-insulin, dexamethasone, indomethacin and IBMX.
2012-12-01 | E-GEOD-37836 | biostudies-arrayexpress
Project description:To investigate the collective role of zinc finger proteins in adipogenesis, we induced adipogenesis using human mesenchymal stem cells (MSC) and compared the gene expression profiles using human Illumina beads arrays. We performed three independent experiments of time-series analysis (0h, 1h, 12h, 24h, 2d, 4d, 6d, 9d, 12d, 15d after induction of adipogenesis) in order to comprehensively capture the dynamic profiles of transcriptome during differentiation. Statistical filtering based on Analysis of Variance (ANOVA; p-value < 0.001, FDR 5%) and Gene Ontology classification using the Human Gene Nomenclature Database (HGNC) identified 618 out of 678 zinc finger proteins (ZFPs) that were significantly expressed during adipogenesis. Moreover, 34 out of 618 genes were differentially expressed more than 2-fold up or down regulated. The K-means clustering analysis further revealed four dynamic expression patterns: one gene in early-response, seven genes in transient, twenty genes in progressively-induced and four genes in the down-regulated, demonstrating the dynamic involvement of ZFPs in the process of adipocyte differentiation and maturation.
2012-12-01 | GSE37836 | GEO
Project description:C2H2 zinc finger proteins represent the largest and most enigmatic class of human transcription factors. Their C2H2 arrays are highly variable, indicating that most will have unique DNA binding motifs. However, most of the binding motifs have not been directly determined. We have determined the binding sites and motifs of 119 C2H2 zinc finger proteins and the expression pattern of 80 cell lines overexpressing C2H2 zinc finger proteins in order to study the role of C2H2 zinc finger proteins in gene regulation. We expressed GFP-tagged C2H2-ZF proteins in stable transgenic HEK293 cells. Total RNA was isolated using Trizol and sequencing libraries were constructed using TruSeq Stranded Total RNA Library Prep Kit with Ribo-Zero Gold or TruSeq RNA Library Preparation Kit v2.
2016-10-25 | E-GEOD-76495 | biostudies-arrayexpress
Project description:Cys2-His2 Zinc finger genes (ZNFs) form the largest family of transcription factors in metazoans. Zebrafish posess a subfamily characterized by the presence of a domain dubbed Fish N-terminal Zinc finger associated (FiNZ). FiNZ-ZNFs are expressed at the onset of zygotic genome activation in zebrafishh, and blocking FiNZ-ZNF translation using morpholinos during early zebrafish embryogenesis results in a broad de-repression of young, transcriptionally active TEs.
2023-04-10 | GSE229157 | GEO
Project description:C2H2 zinc finger proteins represent the largest and most enigmatic class of human transcription factors. Their C2H2 arrays are highly variable, indicating that most will have unique DNA binding motifs. However, most of the binding motifs have not been directly determined. We have determined the binding sites and motifs of 119 C2H2 zinc finger proteins and the expression pattern of 80 cell lines overexpressing C2H2 zinc finger proteins in order to study the role of C2H2 zinc finger proteins in gene regulation.
2016-10-25 | GSE76495 | GEO
Project description:We have identified protein protein interactions of 118 C2H2 zinc finger proteins by AP-MS using GFP-tagged constructs in order to understand their role in transcription regulation.
2016-12-09 | PXD003431 | Pride
Project description:Half of all human transcription factors use C2H2 zinc finger domains to specify site-specific DNA binding and yet very little is known about their role in gene regulation. Based on in vitro studies, a zinc finger code has been developed that predicts a binding motif for a particular zinc finger factor (ZNF). However, very few studies have performed genome-wide analyses of ZNF binding patterns and thus it is not clear if the binding code developed in vitro will be useful for identifying target genes of a particular ZNF. We performed genome-wide ChIP-seq for ZNF263, a C2H2 ZNF that contains 9 finger domains, a KRAB repression domain, and a SCAN domain and identified more than 5000 binding sites in K562 cells. Our results suggest that ZNF263 binds to a 24 nt site which differs from the motif predicted by the zinc finger code in several positions. Interestingly, many of the ZNF263 binding sites are located within the transcribed region of the target gene. Although ZNFs containing a KRAB domain are thought to function mainly as transcriptional repressors, many of the ZNF263 target genes are expressed at high levels. To address the biological role of ZNF263, we identified genes whose expression was altered by treatment of cells with ZNF263-specific siRNAs. Our results suggest that ZNF263 can have both positive and negative effects on transcriptional regulation of its target genes. For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf Examination of ZNF263 ChIP-seq in K562 cells.
2009-12-03 | E-GEOD-19235 | biostudies-arrayexpress