Single base-pair resolution analysis of DNA binding motif with MoMotif reveals an oncogenic function of CTCF zinc-finger 1 mutation.
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PROVIDER: PRJNA785883 | ENA |
REPOSITORIES: ENA
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Project description:This study investigates whether the N-terminal CCHC zinc finger motif is necessary and sufficient for the formation of BCL11B dimers, using fluorescence microscopy and Förster resonance energy transfer-assisted fluorescence-activated cell sorting (FACSFRET). The role of the CCHC motif in homodimer assembly was further investigated using affinity purification followed by mass spectrometry (AP-MS). Functional studies investigated the role of dimerization in the induction of cell cycle arrest and protection against DNA damage-induced apoptosis.
2018-01-12 | PXD008606 | 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.
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Project description:Zinc finger protein SALL4 functions through an AT-rich motif to regulate gene expression
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Project description:Cys2-His2 zinc finger proteins (ZFPs) are the largest group of transcription factors in higher metazoans. A complete characterization of these ZFPs and their associated target sequences is pivotal to fully annotate transcriptional regulatory networks in metazoan genomes. As a first step in this process, we have characterized the DNA-binding specificities of 130 Zinc finger sets from Drosophila melanogaster using a bacterial one-hybrid system. This data set contains the DNA-binding specificities for at least one encoded ZFP from 71 unique genes and 22 alternate splice isoforms. This represents the largest block of characterized ZFPs from any organism described to date. These recognition motifs can be used to predict genomic binding sites and potential regulatory targets for these factors within the fruit fly genome. We have characterized subsets of fingers from these ZFPs to define the correct orientation and register of the zinc fingers on their defined binding sites. By correlating individual fingers with motif subsites, we can assign finger specificity throughout each ZFP. This reveals the diversity of recognition potential within the naturally-occurring zinc fingers of a single organism, where the characterized fingers can specify 47 of the 64 possible DNA triplets. To confirm the utility of our finger recognition models, we have employed subsets of Drosophila fingers in combination with an existing archive of zinc finger modules to create ZFPs with novel DNA-binding specificity. These finger combinations can be used to create novel functional Zinc Finger Nucleases for editing vertebrate genomes. Illumina sequencing of Barcoded Binding sites obtained after B1H selection of Cys2-His2 zinc finger proteins cloned as a C-terminal fusions to the omega subunit of E. coli RNA polymerase in the B1H system.
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Project description:Most cancer associated CTCF mutations are involved in the recognition of CTCF core binding motif. Interestingly, mutations of the currently uncharacterized ZF1 of CTCF are practically exclusive to breast cancer, where they are associated with increased metastatic ability and therapeutic resistance. Although unnecessary to bind the core-binding motif, it is still unknown if the ZF1 of CTCF promotes binding to an altered CTCF motif. Further, classical motif analysis tools are powerless to identify minute changes in core binding motif. Therefore, the biological and epigenetic mechanism of action of CTCF ZF1 mutations is still unexplored. In order to explain the impact of CTCF ZF1M in breast models, we developed a new tool, diffMotif, that allows the identification, at the single base-pair resolution, of extension or alteration of core binding motif. Using diffMotif, we identified an extension of CTCF core binding motif, necessitating a functional ZF1 to bind appropriately. Using a combination of ChIP-Seq and RNA-Seq, we discover that the inability to bind the extended motif led to altered gene expression driving increased invasive ability and drug metabolism, mimicking the harmful oncogenic phenotypes observed clinically. Our study demonstrates the oncologic relevance of the characterisation of mutation induced altered DNA binding ability, by diffMotif.
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Project description:Most cancer associated CTCF mutations are involved in the recognition of CTCF core binding motif. Interestingly, mutations of the currently uncharacterized ZF1 of CTCF are practically exclusive to breast cancer, where they are associated with increased metastatic ability and therapeutic resistance. Although unnecessary to bind the core-binding motif, it is still unknown if the ZF1 of CTCF promotes binding to an altered CTCF motif. Further, classical motif analysis tools are powerless to identify minute changes in core binding motif. Therefore, the biological and epigenetic mechanism of action of CTCF ZF1 mutations is still unexplored. In order to explain the impact of CTCF ZF1M in breast models, we developed a new tool, diffMotif, that allows the identification, at the single base-pair resolution, of extension or alteration of core binding motif. Using diffMotif, we identified an extension of CTCF core binding motif, necessitating a functional ZF1 to bind appropriately. Using a combination of ChIP-Seq and RNA-Seq, we discover that the inability to bind the extended motif led to altered gene expression driving increased invasive ability and drug metabolism, mimicking the harmful oncogenic phenotypes observed clinically. Our study demonstrates the oncologic relevance of the characterisation of mutation induced altered DNA binding ability, by diffMotif.
2022-06-28 | GSE190098 | 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: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.
2009-11-24 | GSE19146 | GEO
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
2009-12-03 | GSE19235 | 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