3′HS1 CTCF binding site in human β-globin locus regulates fetal hemoglobin expression

Abstract

Mutations in the adult β-globin gene can lead to a variety of hemoglobinopathies, including sickle cell disease and β-thalassemia. An increase in fetal hemoglobin expression throughout adulthood, a condition named Hereditary Persistence of Fetal Hemoglobin (HPFH), has been found to ameliorate hemoglobinopathies. Deletional HPFH occurs through the excision of a significant portion of the 3' end of the β-globin locus, including a CTCF binding site termed 3'HS1. Here, we show that the deletion of this CTCF site alone induces fetal hemoglobin expression in both adult CD34+ hematopoietic stem and progenitor cells and HUDEP-2 erythroid progenitor cells. This induction is driven by the ectopic access of a previously postulated distal enhancer located in the OR52A1 gene downstream of the locus, which can also be insulated by the inversion of the 3'HS1 CTCF site. This suggests that genetic editing of this binding site can have therapeutic implications to treat hemoglobinopathies.

Limitations of mouse models for sickle cell disease conferred by their human globin transgene configurations

“…These data indicate that the Townes strain lacks key cis -regulatory elements for controlling human β-globin switching, which could include the long non-coding RNA locus, BGLT3 , the globin pseudogene HBBP1 and the 3′ HS1 downstream of HBB ( Fig. 2 B) ( Himadewi et al, 2021 ; Huang et al, 2017 ; Ivaldi et al, 2018 ). Efforts to model SCD in mice harboring the β-globin YAC have been complicated by excessive perinatal death, most likely because the human γ-to-β-globin gene switch occurs during mid-gestation in mice, thereby eliminating the protective effects of HbF expression during birth-associated hypoxic trauma ( Chang et al, 1998 ).…”

“…The absence of these elements in the Townes strain may impair expression of the HBG1 transgene. In this regard, the HBBP1 and BGLT3 genes ( Huang et al, 2017 ; Ivaldi et al, 2018 ) and 3′ HS1, a CTCF-binding region downstream of HBB ( Himadewi et al, 2021 ), have been shown to regulate γ-globin gene expression, contain human-specific ATAC-seq peaks and are not present in the Townes strain.

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Mutant NPM1 Hijacks Transcriptional Hubs to Maintain Pathogenic Gene Programs in Acute Myeloid Leukemia

“…Chromatin immunoprecipitation was performed using a previously described protocol ( 53 ). Briefly, 1 million cells were fixed with 1% formaldehyde and sonicated with Covaris E220 or Bioruptor Plus.…”

“…mRNA-seq was performed as previously described ( 53 ). Total RNA from OCI-AML3 cells treated with DMSO, dTAG-13, and selinexor (ApexBio, B1464) with specific doses and time was isolated using the Qiagen RNeasy RNA isolation kit (micro).…”

“…Assay for Transposase-Accessible Chromatin using sequencing was performed with the Illumina Nextera XT library preparation kit (FC-131-1024, Illumina) as previously described ( 53 ). Upon 1-day treatment with DMSO, 500 nmol/L dTAG-13, or 25 nmol/L selinexor, 50,000 OCI-AML3 NPM1c degron 2 cells were harvested and permeabilized in 50 μL of cold lysis buffer (10 mmol/L Tris-HCl, pH 7.4, 10 mmol/L NaCl, 3 mmol/L MgCl2, 0.1% IGEPAL CA-630).…”