Epigenome editing of microsatellite repeat enhancers reveals specific regulatory functions and tumor dependencies [NRO-Seq]
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE106915 | GEO | 2018/07/30
REPOSITORIES: GEO
Similar Datasets
Project description:Various types of repetitive sequences are dysregulated in cancer but their contributions to oncogenesis are yet to be determined. Here we combine nascent transcription profiling with epigenome editing to test the function of GGAA microsatellites in Ewing sarcoma, a pediatric bone cancer where the oncogenic fusion protein EWS-FLI1 induces chromatin features of active enhancers at this class of repeats. Silencing of chromatin through epigenome editing at specific GGAA repeats abolished local nascent transcription, decreased the expression of neighboring target genes and, in the case of a repeat associated with the SOX2 locus, reduced the growth of tumor xenografts. Our studies directly demonstrate that repeat elements can acquire enhancer functions in cancer and become critical components of oncogenic gene regulation programs. Raw data for MSC Samples GSM2857574-GSM2857587 are subject to privacy restrictions and were not submitted.
2018-07-30 | GSE106914 | GEO
Project description:Distal enhancers play critical roles in sustaining oncogenic gene expression programs. We obtained ATAC-Seq data and ChIP-Seq data using antibodies against the H3K27ac histone mark, transcription factors ERG and ETV6, and input chromatin from B-ALL cell lines and comparator B-cell cancer cell lines, including B-ALL with biallelic ETV6 inactivation, ETV6-RUNX1, and intact ETV6. We identify aberrant enhancer-like activation of GGAA tandem repeats as a characteristic feature of B-cell acute lymphoblastic leukemia (B-ALL) with genetic defects of the ETV6 transcriptional repressor, including ETV6-RUNX1+ and ETV6-null B-ALL. We show that GGAA repeat enhancers are direct activators of previously identified ETV6-RUNX1+ B-ALL “signature” genes, including likely oncogenic drivers. When restored to ETV6-deficient B-ALL cells, ETV6 directly binds to GGAA repeat enhancers, represses their acetylation, downregulates adjacent genes, and inhibits B-ALL growth. In ETV6-deficient B-ALL cells, we find that the ETS transcription factor ERG directly binds to GGAA microsatellite enhancers and is required for sustained activation of many repeat enhancer-activated genes. Together, our findings reveal a novel epigenetic gatekeeper function of the ETV6 tumor suppressor gene and establish microsatellite enhancers as a key mechanism underlying the unique gene expression program of ETV6-RUNX1+ B-ALL.
2022-10-23 | GSE186936 | GEO
Project description:Distal enhancers play critical roles in sustaining oncogenic gene expression programs. We obtained ATAC-Seq data and ChIP-Seq data using antibodies against the H3K27ac histone mark, transcription factors ERG and ETV6, and input chromatin from B-ALL cell lines and comparator B-cell cancer cell lines, including B-ALL with biallelic ETV6 inactivation, ETV6-RUNX1, and intact ETV6. We identify aberrant enhancer-like activation of GGAA tandem repeats as a characteristic feature of B-cell acute lymphoblastic leukemia (B-ALL) with genetic defects of the ETV6 transcriptional repressor, including ETV6-RUNX1+ and ETV6-null B-ALL. We show that GGAA repeat enhancers are direct activators of previously identified ETV6-RUNX1+ B-ALL “signature” genes, including likely oncogenic drivers. When restored to ETV6-deficient B-ALL cells, ETV6 directly binds to GGAA repeat enhancers, represses their acetylation, downregulates adjacent genes, and inhibits B-ALL growth. In ETV6-deficient B-ALL cells, we find that the ETS transcription factor ERG directly binds to GGAA microsatellite enhancers and is required for sustained activation of many repeat enhancer-activated genes. Together, our findings reveal a novel epigenetic gatekeeper function of the ETV6 tumor suppressor gene and establish microsatellite enhancers as a key mechanism underlying the unique gene expression program of ETV6-RUNX1+ B-ALL.
2022-10-23 | GSE186941 | GEO
Project description:Distal enhancers play critical roles in sustaining oncogenic gene expression programs. We obtained ATAC-Seq data and ChIP-Seq data using antibodies against the H3K27ac histone mark, transcription factors ERG and ETV6, and input chromatin from B-ALL cell lines and comparator B-cell cancer cell lines, including B-ALL with biallelic ETV6 inactivation, ETV6-RUNX1, and intact ETV6. We identify aberrant enhancer-like activation of GGAA tandem repeats as a characteristic feature of B-cell acute lymphoblastic leukemia (B-ALL) with genetic defects of the ETV6 transcriptional repressor, including ETV6-RUNX1+ and ETV6-null B-ALL. We show that GGAA repeat enhancers are direct activators of previously identified ETV6-RUNX1+ B-ALL “signature” genes, including likely oncogenic drivers. When restored to ETV6-deficient B-ALL cells, ETV6 directly binds to GGAA repeat enhancers, represses their acetylation, downregulates adjacent genes, and inhibits B-ALL growth. In ETV6-deficient B-ALL cells, we find that the ETS transcription factor ERG directly binds to GGAA microsatellite enhancers and is required for sustained activation of many repeat enhancer-activated genes. Together, our findings reveal a novel epigenetic gatekeeper function of the ETV6 tumor suppressor gene and establish microsatellite enhancers as a key mechanism underlying the unique gene expression program of ETV6-RUNX1+ B-ALL.
2022-10-23 | GSE186938 | GEO
Project description:Ewing sarcoma usually expresses the EWS/FLI fusion transcription factor oncoprotein. EWS/FLI regulates myriad genes required for Ewing sarcoma development. EWS/FLI binds GGAA-microsatellite sequences in vivo and in vitro, and these sequences provide EWS/FLI-mediated activation to reporter constructs, suggesting that they function as EWS/FLI-response elements. Genomic GGAA-microsatellites are highly variable and polymorphic. Current data suggest that there is an optimal “sweet-spot” GGAA-microsatellite length (of 18-26 GGAA repeats) that confers maximal EWS/FLI-responsiveness to target genes, but the mechanistic basis for this was not known. We now demonstrate the absolute necessity of an EWS/FLI-bound GGAA-microsatellite in regulation of the NR0B1 gene, as well as for Ewing sarcoma proliferation and oncogenic transformation. Biochemical studies, using recombinant Δ22 (a version of EWS/FLI containing only the FLI portion) demonstrated a stoichiometry of one Δ22-monomer binding to every two consecutive GGAA-repeats on shorter GGAA-microsatellite sequences. Surprisingly, the affinity for Δ22 binding to GGAA-microsatellites significantly decreased, and ultimately became unmeasureable, when the size of the GGAA-microsatellite was increased to the “sweet-spot” length. In contrast, a fully-functional EWS/FLI mutant (Mut9) that retains approximately half of the EWS portion of the fusion showed low affinity for smaller GGAA-microsatellites, but instead significantly increased its affinity at “sweet-spot” microsatellite lengths. Single-gene ChIP and genome-wide ChIP-seq and RNA-seq studies extended these findings to the in vivo setting. Taken together, these data demonstrate the absolute requirement of GGAA-microsatellites as EWS/FLI activating response elements in vivo and reveal an unsuspected novel role for the EWS portion of the EWS/FLI fusion in binding to optimal-length GGAA-microsatellites.
2017-10-22 | GSE94502 | GEO
Project description:Ewing sarcoma usually expresses the EWS/FLI fusion transcription factor oncoprotein. EWS/FLI regulates myriad genes required for Ewing sarcoma development. EWS/FLI binds GGAA-microsatellite sequences in vivo and in vitro, and these sequences provide EWS/FLI-mediated activation to reporter constructs, suggesting that they function as EWS/FLI-response elements. Genomic GGAA-microsatellites are highly variable and polymorphic. Current data suggest that there is an optimal “sweet-spot” GGAA-microsatellite length (of 18-26 GGAA repeats) that confers maximal EWS/FLI-responsiveness to target genes, but the mechanistic basis for this was not known. We now demonstrate the absolute necessity of an EWS/FLI-bound GGAA-microsatellite in regulation of the NR0B1 gene, as well as for Ewing sarcoma proliferation and oncogenic transformation. Biochemical studies, using recombinant Δ22 (a version of EWS/FLI containing only the FLI portion) demonstrated a stoichiometry of one Δ22-monomer binding to every two consecutive GGAA-repeats on shorter GGAA-microsatellite sequences. Surprisingly, the affinity for Δ22 binding to GGAA-microsatellites significantly decreased, and ultimately became unmeasureable, when the size of the GGAA-microsatellite was increased to the “sweet-spot” length. In contrast, a fully-functional EWS/FLI mutant (Mut9) that retains approximately half of the EWS portion of the fusion showed low affinity for smaller GGAA-microsatellites, but instead significantly increased its affinity at “sweet-spot” microsatellite lengths. Single-gene ChIP and genome-wide ChIP-seq and RNA-seq studies extended these findings to the in vivo setting. Taken together, these data demonstrate the absolute requirement of GGAA-microsatellites as EWS/FLI activating response elements in vivo and reveal an unsuspected novel role for the EWS portion of the EWS/FLI fusion in binding to optimal-length GGAA-microsatellites.
2017-10-22 | GSE94480 | GEO
Project description:Ewing sarcoma is a bone malignancy of children and young adults, frequently harboring the EWS/FLI t(11;22)(q24;q12) chromosomal translocation. The resulting fusion protein is an aberrant transcription factor that uses highly repetitive GGAA-containing elements (microsatellites) to activate and repress thousands of target genes mediating oncogenesis. However, the mechanisms of EWS/FLI interaction with microsatellites and regulation of target genes expression is not clearly understood. Here, we profile genome-wide protein binding and gene expression. Using a combination of unbiased genome-wide computational and experimental analysis, we define GGAA-microsatellites in a Ewing sarcoma context. Our study identifies two distinct classes of GGAA-microsatellites and demonstrates that EWS/FLI responsiveness is dependent on microsatellite length. At close range (within 5 kb) “promoter-like” microsatellites, EWS/FLI binding and subsequent target genes activation is highly dependent on the number of GGAA-motifs. “Enhancer-like” microsatellites demonstrate a positive correlation with length-dependent EWS/FLI binding, but minimal correlation for activated and none for repressed target genes. Our data suggest that EWS/FLI binds to “promoter-like” and “enhancer-like” microsatellites to mediate activation and repression of target genes through different regulatory mechanisms. Such characterization contributes valuable insight to EWS/FLI transcription factor biology and clarifies the role of GGAA-microsatellites on a global genomic scale. This may provide a unique perspective on the role of non-coding DNA in cancer susceptibility and therapeutic development.
2017-11-09 | GSE99959 | GEO
Project description:Transcription of expanded microsatellite repeats is associated with multiple human diseases, including myotonic dystrophy, Fuchs’ endothelial corneal dystrophy, and C9orf72-ALS/FTD. Eliminating or reducing production of RNA and proteins arising from these expanded loci holds therapeutic benefit. Here, we tested the hypothesis that a deactivated form of the Cas9 enzyme impedes transcription across expanded microsatellites. We observed a repeat length-, PAM-, and strand-dependent reduction in the abundance of repeat-containing RNAs upon targeting dCas9 directly to repeat sequences. Aberrant splicing patterns were rescued in DM1 cells, and production of RAN peptides characteristic of DM1, DM2, and C9orf72-ALS/FTD cells was drastically decreased. Systemic delivery of dCas9/gRNA by adeno-associated virus led to reductions in pathological RNA foci, rescue of chloride channel 1 protein expression, and decreased myotonia. These observations suggest that transcription of microsatellite repeat-containing RNAs is more sensitive to perturbation than transcription of other RNAs, indicating potentially viable strategies for therapeutic intervention.
2017-09-20 | GSE103997 | GEO
Project description:Expansion of the CAG/CTG repeats in functionally unrelated genes is a causative factor in many inherited neurodegenerative disorders, including Huntington's disease (HD), spinocerebellar ataxias (SCAs) and myotonic dystrophy type 1 (DM1). Despite many years of research, the mechanism responsible for repeat instability is unknown, and recent findings indicate the key role of DNA repair in this process. The repair of DSBs induced by genome editing tools results in the shortening of long CAG repeats in yeast models. Understanding this mechanism is the first step to developing a therapeutic strategy based on controlled shortening of repeats. The aim of this study was to characterize Cas9-induced DSB repair products in the endogenous HTT locus in human cells and to identify factors affecting the formation of specific types of mutations. The location of the cleavage site and its sequence affect the outcome of DNA repair. DSBs within CAG repeats result in shortening of the repeats in frame in ~90% of products. The mechanism of this contraction involves MRE11-CTIP and RAD51 activity and extensive DNA end resection reaching ~5000 bp. We demonstrated that a DSB located upstream of CAG repeats induces polymerase theta-mediated end joining, resulting in deletion of the entire CAG tract. Furthermore, using unbiased proteomic analysis, we identified novel factors that may be involved in CAG sequence repair.
2024-11-04 | PXD044960 | Pride
Project description:<p>The Gabriella Miller Kids First Pediatric Research Program (Kids First) is a trans-NIH effort initiated in response to the 2014 Gabriella Miller Kids First Research Act and supported by the NIH Common Fund. This program focuses on gene discovery in pediatric cancers and structural birth defects and the development of the Gabriella Miller Kids First Pediatric Data Resource (Kids First Data Resource).</p> <p>Ewing sarcoma (EWS) is a deadly bone cancer that occurs in children and adolescents. Mounting evidence suggests that a genetic predisposition exists for this pediatric cancer, although the specific genetic contribution has yet to be identified. EWS has never been linked to a specific cancer predisposition syndrome, although several case reports have been published that describe siblings and cousins with EWS. Furthermore, neuroectodermal tumors appear to occur more commonly in families with EWS. The two consistent epidemiology findings in EWS include a very strong Caucasian predilection and increased rates of hernia in EWS patients and their family members. Finally, the role of genetic microsatellite repeats in EWS tumorigenesis has been recently described, and these GGAA microsatellites are polymorphic in repeat size and location across the genome.</p> <p>The study goals of this Kids First project include (1) To identify cancer predisposition genes in EWS trios increasing disease risk, (2) To identify genome-wide GGAA microsatellite repeats in EWS trios increasing disease risk, and (3) To identity de novo mutation and structural variant rates in EWS trios reflecting underlying DNA repair defects that increase disease risk. As part of the Kids First Common Fund initiative, this study proposal will further elucidate the genetic contribution to pediatric cancer development. Around 375 of these trios were selected for whole genome sequencing as part of the Gabriella Miller Kids First fund. The EWS trios have been collected as part of the Children's Oncology Group's AEPI10N5 Study ("Genetic Epidemiology of Ewing Sarcoma"), and each trio has associated phenotypic data including a detailed family history. We will interrogate the sequence data using our genomic analysis pipeline at the University of Utah and the Utah Science Technology and Research initiative's (USTAR) Center for Genetic Discovery. We will look for the genetic contribution to ES and the sequence data with be shared in a repository designated by the Kids First Common Fund.</p> <p>All of the WGS and phenotype data from this study is accessible through <a href="https://kidsfirstdrc.org/" target="_blank">kidsfirstdrc.org</a>, where other Kids First datasets can also be accessed. The WGS of these ~375 EWS trios will help us to understand the genetic origins of a deadly childhood cancer and may lead to novel strategies for prevention and treatment.</p>
| phs001228 | dbGaP