Deletion mapping of regulatory elements for GATA3 in T cells reveals a distal enhancer involved in allergic diseases

Chen, Hsiuyi V.; Lorenzini, Michael H.; Lavalle, Shanna N; Sajeev, Karthyayani; Fonseca, Ariana; Fiaux, Patrick C.; Sen, Arko; Luthra, Ishika; Ho, Aaron J; Chen, Aaron R; Guruvayurappan, Karthik; O’Connor, Carolyn; McVicker, Graham

doi:10.1016/j.ajhg.2023.03.008

Cited by 4 publications

(5 citation statements)

References 46 publications

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“…Several asthma-associated SNPs have been reported to be located in the region around the Gata3 gene body ( 16 – 18 ). By analyzing asthma-associated SNPs from the UK Biobank and GWAS catalog, we confirmed that these SNPs were highly enriched in the region located 926 to 970 kb downstream from GATA3 (referred to as the hG900 region) ( Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Numerous SNPs associated with allergic diseases, autoimmune diseases, and hematopoietic malignancies have been reported near the GATA3 locus. Intriguingly, SNPs associated with hematopoietic malignancies are enriched near the GATA3 gene body ( 18 ). In contrast, SNPs associated with allergic diseases are enriched within the hG900 region ( 18 ) ( Fig.…”

Section: Discussionmentioning

confidence: 99%

“…Intriguingly, SNPs associated with hematopoietic malignancies are enriched near the GATA3 gene body ( 18 ). In contrast, SNPs associated with allergic diseases are enriched within the hG900 region ( 18 ) ( Fig. 1 A ).…”

Section: Discussionmentioning

confidence: 99%

“…Recently, Chen et al have explored functional sequences (FSs) within the entire 2 Mbp surrounding the GATA3 locus using tiling CRISPR screens in Jurkat cells ( 18 ). Notably, 100 bp deletion of seven enhancer regions around FS14, which is located at the center of the hG900 region, demonstrated no impact on GATA3 expression in human Th2 cells.…”

Section: Discussionmentioning

confidence: 99%

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A distal enhancer of GATA3 regulates Th2 differentiation and allergic inflammation

Kumagai,

Iwata,

Furuya

et al. 2024

Proc. Natl. Acad. Sci. U.S.A.

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Asthma is a widespread airway disorder where GATA3-dependent Type-2 helper T (Th2) cells and group 2 innate lymphoid cells (ILC2s) play vital roles. Asthma-associated single nucleotide polymorphisms (SNPs) are enriched in a region located 926-970 kb downstream from GATA3 in the 10p14 (hG900). However, it is unknown how hG900 affects the pathogenesis of allergic airway inflammation. To investigate the roles of the asthma-associated GATA3 enhancer region in experimental allergic airway inflammation, we first examined the correlation between GATA3 expression and the activation of the hG900 region was analyzed by flow cytometry and ChIP-qPCR. We found that The activation of enhancers in the hG900 region was strongly correlated to the levels of GATA3 in human peripheral T cell subsets. We next generated mice lacking the mG900 region (mG900KO mice) were generated by the CRISPR-Cas9 system, and the development and function of helper T cells and ILCs in mG900KO mice were analyzed in steady-state conditions and allergic airway inflammation induced by papain or house dust mite (HDM). The deletion of the mG900 did not affect the development of lymphocytes in steady-state conditions or allergic airway inflammation induced by papain. However, mG900KO mice exhibited reduced allergic inflammation and Th2 differentiation in the HDM-induced allergic airway inflammation. The analysis of the chromatin conformation around Gata3 by circular chromosome conformation capture coupled to high-throughput sequencing (4C-seq) revealed that the mG900 region interacted with the transcription start site of Gata3 with an influencing chromatin conformation in Th2 cells. These findings indicate that the mG900 region plays a pivotal role in Th2 differentiation and thus enhances allergic airway inflammation.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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A distal enhancer of GATA3 regulates Th2 differentiation and allergic inflammation

Kumagai,

Iwata,

Furuya

et al. 2024

Proc. Natl. Acad. Sci. U.S.A.

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“…The gRNA pairs included the following: NMU_tss+NMU_tss (positive control), Safe_harbor (SH)+SH (negative control), A_sgRNA1+SH, A_sgRNA2+SH, SH+B_sgRNA1, SH+B_sgRNA2, A_sgRNA1+B_sgRNA1, A_sgRNA1+B_sgRNA2, A_sgRNA2+B_sgRNA1, A_sgRNA2+B_sgRNA2. Pairs of gRNAs were cloned into pLV-dCas9-KRAB-puro (Addgene #71236) following published methods 33,34 . Briefly, DNA oligos carrying pairs of guides were synthesized by IDT and cloned into pLV-dCas9-KRAB-puro plasmids by Gibson assembly reactions.…”

Section: Crispri Perturbation Of Nmu Enhancersmentioning

confidence: 99%

Analysis of single-cell CRISPR perturbations indicates that enhancers act multiplicatively and provides limited evidence for epistatic-like interactions

Zhou

Guruvayurappan

Chen

et al. 2023

Preprint

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A single gene may be regulated by multiple enhancers, but how they work in concert to regulate transcription is poorly understood. Prior studies have mostly examined enhancers at single loci and have reached inconsistent conclusions about whether epistatic-like interactions exist between them. To analyze enhancer interactions throughout the genome, we developed a statistical framework for CRISPR regulatory screens that utilizes negative binomial generalized linear models that account for variable guide RNA (gRNA) efficiency. We reanalyzed a single-cell CRISPR interference experiment that delivered random combinations of enhancer-targeting gRNAs to each cell and interrogated interactions between 3,808 enhancer pairs. We found that enhancers act multiplicatively with one another to control gene expression, but our analysis provides no evidence for interaction effects between pairs of enhancers regulating the same gene. Our findings illuminate the regulatory behavior of multiple enhancers and our statistical framework provides utility for future analyses studying interactions between enhancers.

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DNA-binding factor footprints and enhancer RNAs identify functional non-coding genetic variants

Biddie,

Weykopf,

Hird

et al. 2023

Preprint

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Genome-wide association studies (GWAS) have revealed a multitude of candidate genetic variants affecting the risk of developing complex traits and diseases. However, these highlighted regions are typically in the non-coding genome, and uncovering the functional causative single nucleotide variants (SNVs) is challenging. Prioritisation of variants is commonly based on functional genomic annotation with markers of active regulatory elements, but current approaches still poorly predict functional variants. To address this, we systematically analyse six markers of active regulatory elements for their ability to identify functional variants. We benchmark against molecular quantitative trait loci (molQTL) from assays of regulatory element activity that identify allelic effects on DNA-binding factor occupancy, reporter assay expression, and chromatin accessibility. We identify the combination of DNase footprints and divergent enhancer RNA as markers for functional variants. This signature provides high precision, trading-off low recall, thus substantially reducing candidate variant sets to prioritise variants for functional validation. We present this as a framework called FINDER – Functional SNV IdeNtification using DNase footprints and Enhancer RNA, and demonstrate its utility to prioritise variants using leukocyte count trait and analyse variants in linkage disequilibrium with a lead variant to predict a functional variant in asthma. Our findings have implications for prioritising variants from GWAS, in development of predictive scoring algorithms, and for functionally informed fine mapping approaches.

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Deletion mapping of regulatory elements for GATA3 in T cells reveals a distal enhancer involved in allergic diseases

Cited by 4 publications

References 46 publications

A distal enhancer of GATA3 regulates Th2 differentiation and allergic inflammation

A distal enhancer of GATA3 regulates Th2 differentiation and allergic inflammation

Analysis of single-cell CRISPR perturbations indicates that enhancers act multiplicatively and provides limited evidence for epistatic-like interactions

DNA-binding factor footprints and enhancer RNAs identify functional non-coding genetic variants

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