Single-nucleotide-level mapping of DNA regulatory elements that control fetal hemoglobin expression

Cheng, Li; Li, Yichao; Qi, Qian; Peng, Xu; Feng, Ruopeng; Palmer, Lance E.; Chen, Jingjing; Wu, Ruiqiong; Yee, Tiffany; Zhang, Jingjing; Yao, Yu; Sharma, Akshay; Hardison, R; Weiss, Mitchell J.; Cheng, Yong

doi:10.1038/s41588-021-00861-8

Cited by 47 publications

(34 citation statements)

References 92 publications

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“…This will ultimately allow tackling polygenic traits in the organismal context. It will likewise provide experimental access to the regulatory genome, as recently demonstrated by combining epigenomics with adenine base editing in vitro that mapped and validated sickle cell disease-associated cis-regulatory elements of the fetal haemoglobin (Cheng et al, 2021). Modeling SNVs mutations may be achieved with stringent criteria (no bystander mutations accepted, n=1951) or less stringent selection (allowing bystander mutations "all", n=5896).…”

Section: Discussionmentioning

confidence: 99%

Precise in vivo functional analysis of DNA variants with base editing using ACEofBASEs target prediction

Cornean

Gierten

Welz

et al. 2021

Preprint

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Single nucleotide variants (SNVs) are prevalent genetic factors shaping individual trait profiles and disease susceptibility. The recent development and optimizations of base editors, rubber and pencil genome editing tools now promise to enable direct functional assessment of SNVs in model organisms. However, the lack of bioinformatic tools aiding target prediction limits the application of base editing in vivo>. Here, we provide a framework for adenine and cytosine base editing in medaka (Oryzias latipes) and zebrafish (Danio rerio), ideal for scalable validation studies. We developed an online base editing tool ACEofBASEs (a careful evaluation of base-edits), to facilitate decision-making by streamlining sgRNA design and performing off-target evaluation. We used state-of-the-art adenine (ABE) and cytosine base editors (CBE) in medaka and zebrafish to edit eye pigmentation genes and transgenic GFP function with high efficiencies. Base editing in the genes encoding troponin T and the potassium channel ERG faithfully recreated known cardiac phenotypes. We finally validated missense mutations in novel candidate genes of congenital heart disease (CHD) dapk3, ube2b, usp44, and ptpn11 and present the resulting heart specific phenotypes. This base editing framework applies to a wide range of SNV-susceptible traits accessible in fish, facilitating straight-forward candidate validation and prioritization for detailed mechanistic downstream studies.

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Section: Discussionmentioning

confidence: 99%

Precise in vivo functional analysis of DNA variants with base editing using ACEofBASEs target prediction

Cornean

Gierten

Welz

et al. 2021

Preprint

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show abstract

“…A recent study applied an APOBECB-mediated base editor screen to map regulatory elements of four loci involved in HbF expression. A total of 6,174 sgRNAs targeting 307 putative regulatory elements were investigated by correlating enriched sgRNAs to HbF expression abundance, revealing novel therapeutic candidates for sickle cell disease treatment (Cheng et al, 2021).…”

Section: Mapping Regulatory Elementsmentioning

confidence: 99%

Gene editing and its applications in biomedicine

Zhuang

et al. 2022

Sci. China Life Sci.

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The steady progress in genome editing, especially genome editing based on the use of clustered regularly interspaced short palindromic repeats (CRISPR) and programmable nucleases to make precise modifications to genetic material, has provided enormous opportunities to advance biomedical research and promote human health. The application of these technologies in basic biomedical research has yielded significant advances in identifying and studying key molecular targets relevant to human diseases and their treatment. The clinical translation of genome editing techniques offers unprecedented biomedical engineering capabilities in the diagnosis, prevention, and treatment of disease or disability. Here, we provide a general summary of emerging biomedical applications of genome editing, including open challenges. We also summarize the tools of genome editing and the insights derived from their applications, hoping to accelerate new discoveries and therapies in biomedicine.

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“…The shaded areas highlighting the HBG1 and HBG2 promoters are shown in higher resolution below. b , ChIP seq analysis for GATA1 in HUDEP-1 and HUDEP-2 cells, which express predominantly γ-globin and β-globin respectively, shown as described for panel a. GATA1 occupancy in HUDEP-2 cells was derived from publicly available data 82 .…”

Section: Extended Datamentioning

confidence: 99%

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

Doerfler

Feng

et al. 2021

Nat Genet

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Hereditary persistence of fetal hemoglobin (HPFH) ameliorates β-hemoglobinopathies by inhibiting the developmental switch from γ-globin ( HBG1 / HBG2 ) to β-globin ( HBB ) gene expression. Some forms of HPFH are associated with γ-globin promoter variants that either disrupt binding motifs for transcriptional repressors or create new motifs for transcriptional activators. How these variants sustain γ-globin gene expression postnatally remains undefined. We mapped γ-globin promoter sequences functionally in erythroid cells harboring different HPFH variants. Those that disrupt a BCL11A repressor binding element induce γ-globin expression by facilitating the recruitment of transcription factors NF-Y to a nearby proximal CCAAT box and GATA1 to an upstream motif. The proximal CCAAT element becomes dispensable for HPFH variants that generate new binding motifs for activators NF-Y or KLF1, but GATA1 recruitment remains essential. Our findings define distinct mechanisms through which transcription factors and their cis- regulatory elements activate γ-globin expression in different forms of HPFH, some of which are being recreated by therapeutic genome editing.

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Single-nucleotide-level mapping of DNA regulatory elements that control fetal hemoglobin expression

Cited by 47 publications

References 92 publications

Precise in vivo functional analysis of DNA variants with base editing using ACEofBASEs target prediction

Precise in vivo functional analysis of DNA variants with base editing using ACEofBASEs target prediction

Gene editing and its applications in biomedicine

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

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