Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Micro-Capture-C to determine the long-range interaction of HBG1/2 promoter with the locus control region (LCR), a powerful upstream enhancer in -175 A>G mutantant HUDEP-2 cells.

ABSTRACT: Naturally occurring mutations in the γ-globin promoters can result in hereditary persistence of fetal hemoglobin (HPFH), a benign condition that ameliorates the severity of β-hemoglobinopathies through increased post-natal fetal hemoglobin (HbF) expression. Base editing to recreate the HPFH mutations is a promising approach to induce HbF. Compared with Cas9-generated indels, base-editing approaches were more potent, with the −175 A>G conversion resulting in the strongest induction of HbF by creating a new TAL1 binding motif that stimulates long-range interaction with the locus control region (LCR), a powerful upstream enhancer. Micro-Capture-C analysis showed that introducing the −175 A>G variant stimulated long-range interaction between the γ-globin promoters and the LCR in erythroid progenitor cell line. Together, these findings show that the −175 A>G variant creates a bipartite GATA–TAL1 motif, resulting in the assembly of a GATA1/TAL1/LMO2/LDB1 complex, which activates γ-globin gene transcription by enhancing its physical association with the LCR.

ORGANISM(S): Homo sapiens

PROVIDER: GSE228570 | GEO | 2023/04/28

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Potent and uniform fetal hemoglobin induction via base editing [Cut & Run]

Project description:Genetic manipulations to increase fetal hemoglobin (HbF, α2γ2) in postnatal red blood cells (RBCs) can alleviate β-thalassemia and sickle cell disease. We compared five strategies in CD34+ hematopoietic stem and progenitor cells, using either Cas9 nuclease, which creates uncontrolled indel mixtures, or adenine base editors (ABE), which generate more precise nucleotide changes. The most potent modification was ABE generation of γ-globin (HBG1/HBG2) −175 A>G, which creates a promoter binding motif for the transcriptional activator TAL1. In erythroid colonies with >87.5% on-target edits, those with −175 A>G expressed 81 ± 7% HbF, versus 17 ± 11% in unedited controls. In comparison, HbF levels were lower and more variable in erythroid cells modified with either of two Cas9 nuclease strategies with similar editing efficiencies, being 32 ± 19% when a BCL11A repressor binding motif in the γ-globin promoter was targeted and 52 ± 13% when the +58 BCL11A erythroid enhancer was targeted. Contrary to currently accepted models of γ-globin regulation, HbF levels varied significantly with different Cas9 indels that disrupted the γ-globin promoter BCL11A binding motif. The −175 A>G base edit also induced HbF more potently than did the Cas9 nuclease approaches in RBCs generated after transplantation of modified normal or SCD patient CD34+ cells into mice. Our data suggest a strategy for potent, uniform induction of HbF and provide insights into γ-globin gene regulation. More generally, we demonstrate that diverse indels generated by Cas9 nuclease can cause unexpected variations in biological outcomes that can be circumvented by base editing, with important implications for therapeutic gene editing efforts.

2023-04-28 | GSE228819 | GEO

CIRCLE-seq analyais for -175 site

Project description:Adenosine base editing to recreate the HPFH mutations is a promising approach to induce HbF. We used CIRCLE-seq to identify 682 potential off-target sites in purified DNA treated with Cas9 nuclease complexed to sgRNA targeting -globin -175 .

2023-04-28 | GSE228569 | GEO

Editing the core region in HPFH deletions alters fetal and adult globin expression for treatment of β-hemoglobinopathies [4C]

Project description:We have identified putative repressor region (PRR) to exon-1 of β-globin (βE1) in the β-globin cluster as a core region in alll the HPFH deletions that activates fetal hemoglobin and silences adult hemoglobin. To test the impact of PRR-βE1 gene editing in β-globin cluster configuration, we employed Circular chromosome conformation capture (4C) analysis to assess the LCR interaction to the gamma-globin promoter. Using HBG2 promoter as viewpoint,4C analysis showed an increased interaction of HBG promoter to the hypersensitive (HS) sites of the LCR in HUDEP-2 clones harbouring PRR-βE1 biallelic deletions compared to control HUDEP-2 cells. Link for 4C output files Mendeley - Venkatesan, Vigneshwaran (2022), “4C data set ”, Mendeley Data, V1, doi: 10.17632/cz3thnr9kf.1

2022-04-26 | GSE201388 | GEO

Transcriptome-wide deamination analysis

Project description:Adenosine base editing to recreate the HPFH mutations is a promising approach to induce HbF. To measure Cas-independent off-target editing of mRNA, we electroporated CD34+ cells from three different donors with RNP consisting of ABE7.10 or ABE8e complexed with the -globin −175 A>G targeting sgRNA. Six days later, this was followed by high-depth RNA-seq. The rates of A-to-I conversion were similar in edited and unedited control cells, which is consistent with previous reports that off-target RNA editing was not detected after high-level adenine base editing of HUDEP2 cells or HSPCs.

2023-04-28 | GSE228821 | GEO

Transcription factor competition at the γ-globin promoters controls hemoglobin switching [CRISPR Screen]

Project description:BCL11A, the major regulator of HbF(α2γ2) level, represses γ-globin expression through direct promoter binding in adult erythroid cells in a switch to adult adult-type HbA (α2β2) production. Yet, the mechanism remains unclear. To uncover how BCL11A initiates repression, we used CRISPR/Cas9 and dCas9 screens to dissect the γ-globin promoters and identified an apparent activator element near the BCL11A binding region. Using CUT&RUN and base editing approaches, we demonstrate that this element, the proximal CCAAT box, is the binding site of transcription activator NF-Y. BCL11A competes with NF-Y binding through steric hindrance to initiate γ-globin repression, and the distance between the two motifs is critical for direct competition. Occupancy of NF-Y is rapidly established upon BCL11A depletion, and precedes γ-globin derepression and LCR-globin loop formation. Our findings reveal that the critical fetal-to-adult hemoglobin switch is initiated by the competition between transcription factors within a discrete region in the γ-globin promoters.

2020-12-22 | GSE160456 | GEO

Transcription factor competition at the γ-globin promoters controls hemoglobin switching [ChIP-seq]

2020-12-22 | GSE150490 | GEO

Transcription factor competition at the γ-globin promoters controls hemoglobin switching [ATACseq]

2020-12-22 | GSE150488 | GEO

Transcription factor competition at the γ-globin promoters controls hemoglobin switching [CUT&RUN]

2020-12-22 | GSE150529 | GEO

Transcription factor competition at the γ-globin promoters controls hemoglobin switching [PRO-seq]

2020-12-22 | GSE150491 | GEO

Activation of γ-globin gene expression by GATA1 and NF-Y in hereditary persistence of fetal hemoglobin

Project description:Naturally occurring mutations in the Î³-globin promoters can result in hereditary persistence of fetal hemoglobin (HPFH), a benign condition that ameliorates the severity of Î²-hemoglobinopathies through increased post-natal fetal hemoglobin (HbF) expression. Mutations in the Î³-globin promoters result in loss or de novo recruitment of transcription factors, yet the cis-regulatory elements involved in Î³-globin transcription in the context of HPFH are not clearly defined. We demonstrate that the -115 cluster of HPFH mutations require GATA1 binding to at -186 in cooperation with NF-Y at the proximal CCAAT box at -85 for Î³-globin expression. We show that multiple HPFH mutations increase HbF in an erythroid cell line and result in GATA1 binding at -186. Mutation of the -186 GATA motif in HPFH erythroid cell lines and erythroid-differentiated CD34+ cells resulted in loss of GATA1 binding and reduction in fetal hemoglobin. NF-Y ChIP-seq in HPFH erythroid cell lines demonstrated binding to the proximal CCAAT box at -85 was also found to be critical for HPFH-mediated Î³-globin expression. Together, the -186 GATA motif and -85 proximal CCAAT box function in an additive and independent manner. Our results reveal detailed evidence for common, positive acting cis-regulatory elements that may provide more general mechanisms for erythroid gene expression.

2021-04-22 | GSE152338 | GEO

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data