TRIM28 and novel interacting KRAB-ZNFs preserve self-renewal of human pluripotent stem cells through H3K9me3 and DNA methylation mediated repression of pro-differentiation genes
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE97403 | GEO | 2017/11/30
SECONDARY ACCESSION(S): PRJNA381641
REPOSITORIES: GEO
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Project description:Reprogramming to induced pluripotent stem cells (iPSCs) and differentiation of pluripotent stem cells (PSCs) are primarily regulated by epigenetic machinery. Tripartite motif protein 28 (TRIM28), a universal mediator of Krüppel-associated box domain zinc fingers (KRAB-ZNFs), is known to regulate both processes, however exact mechanism and identity of participating KRAB-ZNF genes remains unknown. Here, using a novel reporter system, we first showed that TRIM28/KRAB-ZNFs alter DNA methylation patterns in addition to H3K9me3 to cause gene repression during reprogramming. Next, using several expression datasets, we identified novel KRAB-ZNFs (ZNF114, ZNF483 and ZNF589) in the human genome that regulate pluripotency. Mechanistically, we demonstrated that these KRAB-ZNFs directly alter gene expression of important developmental genes by modulation of H3K9me3 and DNA methylation on their promoters. In summary, TRIM28 employs novel KRAB-ZNFs to evoke epigenetic silencing of its target differentiation genes by H3K9me3 and DNA methylation.
2018-01-10 | GSE95096 | GEO
Project description:Only a small percentage of human transcription factors (e.g. those associated with a specific differentiation program) are expressed in a given cell type. Thus, cell fate is mainly determined by cell type-specific silencing of transcription factors that drive different cellular lineages. Several histone modifications have been associated with gene silencing, including H3K27me3 and H3K9me3. We have previously shown that the two largest classes of mammalian transcription factors are marked by distinct histone modifications; homeobox genes are marked by H3K27me3 and zinc finger genes are marked by H3K9me3. Several histone methyltransferases (e.g. G9a and SETDB1) may be involved in mediating the H3K9me3 silencing mark. We have used ChIP-chip and ChIP-seq (GSE24632) to demonstrate that SETDB1, but not G9a, is associated with regions of the genome enriched for H3K9me3. A current model is that SETDB1 is recruited to specific genomic locations via interaction with the corepressor TRIM28 (KAP1), which is in turn recruited to the genome via interaction with zinc finger transcription factors that contain a Kruppel-associated box (KRAB) domain. However, specific KRAB-ZNFs that recruit TRIM28 (KAP1) and SETDB1 to the genome have not been identified. We now show that ZNF274 (a KRAB-ZNF that contains 5 C2H2 zinc finger domains), can interact with KAP1 in vitro and, using ChIP-seq, we show that ZNF274 binding sites co-localize with SETDB1, KAP1, and H3K9me3 at the 3â ends of zinc finger genes. Knockdown of ZNF274 with siRNAs reduced the levels of KAP1 and SETDB1 recruitment to the binding sites. These studies provide the first identification of a KRAB domain-containing ZNFs that is involved in recruitment of the KAP1 and SETDB1 to the human genome. This study includes the 4 ChIP-chip arrays only.
2010-11-01 | E-GEOD-24480 | biostudies-arrayexpress
Project description:Only a small percentage of human transcription factors (e.g. those associated with a specific differentiation program) are expressed in a given cell type. Thus, cell fate is mainly determined by cell type-specific silencing of transcription factors that drive different cellular lineages. Several histone modifications have been associated with gene silencing, including H3K27me3 and H3K9me3. We have previously shown that the two largest classes of mammalian transcription factors are marked by distinct histone modifications; homeobox genes are marked by H3K27me3 and zinc finger genes are marked by H3K9me3. Several histone methyltransferases (e.g. G9a and SETDB1) may be involved in mediating the H3K9me3 silencing mark. We have used ChIP-chip (GSE24480) and ChIP-seq to demonstrate that SETDB1, but not G9a, is associated with regions of the genome enriched for H3K9me3. A current model is that SETDB1 is recruited to specific genomic locations via interaction with the corepressor TRIM28 (KAP1), which is in turn recruited to the genome via interaction with zinc finger transcription factors that contain a Kruppel-associated box (KRAB) domain. However, specific KRAB-ZNFs that recruit TRIM28 (KAP1) and SETDB1 to the genome have not been identified. We now show that ZNF274 (a KRAB-ZNF that contains 5 C2H2 zinc finger domains), can interact with KAP1 in vitro and, using ChIP-seq, we show that ZNF274 binding sites co-localize with SETDB1, KAP1, and H3K9me3 at the 3’ ends of zinc finger genes. Knockdown of ZNF274 with siRNAs reduced the levels of KAP1 and SETDB1 recruitment to the binding sites. These studies provide the first identification of a KRAB domain-containing ZNFs that is involved in recruitment of the KAP1 and SETDB1 to the human genome. For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf 7 total ChIP-seq datasets; 4 ZNF274 datasets done in duplicate from 4 different cell lines; 1 KAP1 duplicate dataset done in duplicate from K562 cells; 1 SetDB1 duplicate dataset from K562 cells; 1 H3K9me3 duplicate dataset from K562 cells
2010-11-15 | E-GEOD-24632 | biostudies-arrayexpress
Project description:Since the discovery of induced pluripotent stem cells there has been intense interest in understanding the mechanisms that allow a somatic cell to be reprogrammed back to a pluripotent state. Several groups have studied the alterations in gene expression that occur as somatic cells modify their genome to that of an embryonic stem cell. Underpinning many of the gene expression changes are modifications to the epigenetic profile of the associated chromatin. We have used a large-scale shRNA screen to identify epigenetic modifiers that act as barriers to reprogramming. We have uncovered an important role for TRIM28 in cells resisting transition between somatic and pluripotent states. TRIM28 achieves this by maintaining the H3K9me3 repressed state and keeping endogenous retroviruses silenced. We propose that knockdown of TRIM28 during reprogramming results in more plastic H3K9me3 domains, dysregulation of genes nearby H3K9me3 marks, and up regulation of endogenous retroviruses, thus facilitating the transition through reprogramming. Gene expression profiling using high through put sequencing at day 7 of Oct4, Sox2, Klf4 and cMyc (OSKM) expression in mouse embryonic fibroblasts with or without Trim28 / Setdb1 knockdown
2016-06-28 | E-GEOD-80550 | biostudies-arrayexpress
Project description:Only a small percentage of human transcription factors (e.g. those associated with a specific differentiation program) are expressed in a given cell type. Thus, cell fate is mainly determined by cell type-specific silencing of transcription factors that drive different cellular lineages. Several histone modifications have been associated with gene silencing, including H3K27me3 and H3K9me3. We have previously shown that the two largest classes of mammalian transcription factors are marked by distinct histone modifications; homeobox genes are marked by H3K27me3 and zinc finger genes are marked by H3K9me3. Several histone methyltransferases (e.g. G9a and SETDB1) may be involved in mediating the H3K9me3 silencing mark. We have used ChIP-chip and ChIP-seq (GSE24632) to demonstrate that SETDB1, but not G9a, is associated with regions of the genome enriched for H3K9me3. A current model is that SETDB1 is recruited to specific genomic locations via interaction with the corepressor TRIM28 (KAP1), which is in turn recruited to the genome via interaction with zinc finger transcription factors that contain a Kruppel-associated box (KRAB) domain. However, specific KRAB-ZNFs that recruit TRIM28 (KAP1) and SETDB1 to the genome have not been identified. We now show that ZNF274 (a KRAB-ZNF that contains 5 C2H2 zinc finger domains), can interact with KAP1 in vitro and, using ChIP-seq, we show that ZNF274 binding sites co-localize with SETDB1, KAP1, and H3K9me3 at the 3’ ends of zinc finger genes. Knockdown of ZNF274 with siRNAs reduced the levels of KAP1 and SETDB1 recruitment to the binding sites. These studies provide the first identification of a KRAB domain-containing ZNFs that is involved in recruitment of the KAP1 and SETDB1 to the human genome.
2010-11-01 | GSE24480 | GEO
Project description:KAP1 is overexpressed in breast cancer. To determine KAP1 regulated genes, we performed microarray analysis of gene expression in KAP1 depleted breast cancer cells MDA-MB-231LN. The transcriptional regulator TRIM28/KAP1 plays an important role in development, stem cell self-renewal, chromatin organization and the DNA damage response. KAP1 is an essential co-repressor for KRAB zinc finger proteins (KRAB-ZNFs). Though KRAB-ZNFs represent the largest family of human transcription factors, their biological functions are largely unknown. Using the conserved zinc fingers linker region (ZnFL) as antigen, we have developed a ZnFL antibody that recognizes multiple KRAB-ZNFs. We showed that KAP1 and many KRAB-ZNFs were overexpressed in human breast cancers and breast cancer cell lines. In addition, an active SUMOylated form of KAP1 was markedly increased in breast cancer cells. Furthermore, KAP1 depletion in breast cancer cell lines reduced cell proliferation and inhibited tumor growth and metastasis of tumor xenografts. Conversely, KAP1 overexpression stimulated cell proliferation and tumor growth. KAP1 knockdown led to down-regulation of genes previously linked to tumor progression and metastasis, including PTGS2/COX2, EREG, CD44, MMP1 and MMP2. Interestingly, KAP1 depletion or genomic deletion led to dramatic down-regulation of multiple KRAB-ZNF proteins due in part to their increased degradation. KAP1-dependent stabilization of KRAB-ZNFs required a direct KRAB-ZNF-KAP1 interaction. These results establish KAP1 as a positive regulator of multiple KRAB-ZNFs and an important factor in the development of breast cancer. 7 total samples were analyzed. Stable sublines of MDA-MB-231LN cells expressing control non-targeting shRNA (Scr, 3 biological replicates) and two different shRNAs against KAP1 (KAP1-3, 2 biological replicates and KAP1-4, 2 biological replicates) from doxycycline-inducible pTRIPZ vector were cultured in the presence of 0.5 ug/ml doxycycline for 7 days to induce shRNA expression. Cells were lysed and total RNA was isolated using mirVana miRNA isolation kit (Ambion) in the WVU Genomics Core Facility.
2014-09-23 | E-GEOD-61639 | biostudies-arrayexpress
Project description:Only a small percentage of human transcription factors (e.g. those associated with a specific differentiation program) are expressed in a given cell type. Thus, cell fate is mainly determined by cell type-specific silencing of transcription factors that drive different cellular lineages. Several histone modifications have been associated with gene silencing, including H3K27me3 and H3K9me3. We have previously shown that the two largest classes of mammalian transcription factors are marked by distinct histone modifications; homeobox genes are marked by H3K27me3 and zinc finger genes are marked by H3K9me3. Several histone methyltransferases (e.g. G9a and SETDB1) may be involved in mediating the H3K9me3 silencing mark. We have used ChIP-chip (GSE24480) and ChIP-seq to demonstrate that SETDB1, but not G9a, is associated with regions of the genome enriched for H3K9me3. A current model is that SETDB1 is recruited to specific genomic locations via interaction with the corepressor TRIM28 (KAP1), which is in turn recruited to the genome via interaction with zinc finger transcription factors that contain a Kruppel-associated box (KRAB) domain. However, specific KRAB-ZNFs that recruit TRIM28 (KAP1) and SETDB1 to the genome have not been identified. We now show that ZNF274 (a KRAB-ZNF that contains 5 C2H2 zinc finger domains), can interact with KAP1 in vitro and, using ChIP-seq, we show that ZNF274 binding sites co-localize with SETDB1, KAP1, and H3K9me3 at the 3’ ends of zinc finger genes. Knockdown of ZNF274 with siRNAs reduced the levels of KAP1 and SETDB1 recruitment to the binding sites. These studies provide the first identification of a KRAB domain-containing ZNFs that is involved in recruitment of the KAP1 and SETDB1 to the human genome. For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf
2010-11-15 | GSE24632 | GEO
Project description:TRIM28 and novel interacting KRAB-ZNFs preserve self-renewal of human pluripotent stem cells through H3K9me3 and DNA methylation mediated repression of pro-differentiation genes
| PRJNA381641 | ENA
Project description:Here, we report that ATRX co-localizes with the H3K9-methyl transferase SETDB1 (also known as ESET), the co-repressor TRIM28 (also known as KAP1), and the transcription factor ZNF274 at 3â exons of Zinc Finger Genes (ZNFs) containing an atypical H3K9me3/H3K36me3 chromatin signature. Disruption of ATRX and ZNF274 leads to a significant reduction of H3K9me3, particularly at the 3â ZNF exons and other atypical chromatin regions, higher percentages of DNA damage, and defects in cell cycle. Taken together, our studies suggest that ATRX binds the 3â exons of ZNFs to maintain genomic stability through the regulation of their H3K9me3 levels XL-MNase ChIP-seq of ATRX was performed in the erythroleukemic cell line K562 and the Neuroblastoma cell line LAN6. Two independent replicates using different ATRX antibodies were performed in K562. Additionally, Native ChIP-seq of H3K9me3 in LAN6, ATRX WT and ATRX KO K562 cells was performed. Input samples were sequenced as control.
2016-04-05 | E-GEOD-70920 | biostudies-arrayexpress
Project description:Here, we report that ATRX co-localizes with the H3K9-methyl transferase SETDB1 (also known as ESET), the co-repressor TRIM28 (also known as KAP1), and the transcription factor ZNF274 at 3’ exons of Zinc Finger Genes (ZNFs) containing an atypical H3K9me3/H3K36me3 chromatin signature. Disruption of ATRX and ZNF274 leads to a significant reduction of H3K9me3, particularly at the 3’ ZNF exons and other atypical chromatin regions, higher percentages of DNA damage, and defects in cell cycle. Taken together, our studies suggest that ATRX binds the 3’ exons of ZNFs to maintain genomic stability through the regulation of their H3K9me3 levels
2016-04-05 | GSE70920 | GEO