Pathogenic BCL11A variants provide insights into the mechanisms of human fetal hemoglobin silencing

Abstract

Increased production of fetal hemoglobin (HbF) can ameliorate the severity of sickle cell disease and β-thalassemia. BCL11A has been identified as a key regulator of HbF silencing, although its precise mechanisms of action remain incompletely understood. Recent studies have identified pathogenic mutations that cause heterozygous loss-of-function of BCL11A and result in a distinct neurodevelopmental disorder that is characterized by persistent HbF expression. While the majority of cases have deletions or null mutations causing haploinsufficiency of BCL11A, several missense variants have also been identified. Here, we perform functional studies on these variants to uncover specific liabilities for BCL11A’s function in HbF silencing. We find several mutations in an N-terminal C2HC zinc finger that increase proteasomal degradation of BCL11A. We also identify a distinct C-terminal missense variant in the fifth zinc finger domain that we demonstrate causes loss-of-function through disruption of DNA binding. Our analysis of missense variants causing loss-of-function in vivo illuminates mechanisms by which BCL11A silences HbF and also suggests potential therapeutic avenues for HbF induction to treat sickle cell disease and β-thalassemia.

NMR structure verifies the eponymous zinc finger domain of transcription factor ZNF750

“…We think it is useful to distinguish this knuckle-CCHC family from the finger-CCHC family exemplified by the Z* domain, since the latter have structures very similar to the classical ZNFs. Besides Z*, other examples of finger-CCHC domains exist in the proteins NEMO ( Cordier et al, 2008 ), Fog, MOZ, U-shaped ( Matthews et al, 2000 ), and BLC11A ( Grabarczyk et al, 2018 , Shen et al, 2021 ).…”

Genetic regulation of fetal hemoglobin across global populations

“…Lentiviral particles were produced as previously described. 8 Briefly, 293T cells cultured in DMEM supplemented with 10% FBS were co-transfected with packaging vectors pVSVG and pΔ8.9, and the expression vectors HMD-empty vector or HMD-BACH2. DMEM was replaced with erythroid differentiation base media 24 h later and supernatant containing lentivirus were collected, filtered with a 0.45 μm filter, and concentrated by ultracentrifugation (24,000 rpm, 2 h, 4 0 C).…”

“…The frequency of F-cells in transduced HSPCs undergoing erythroid differentiation was quantified as previously described. 8 Briefly, cells were fixed in 0.05% glutaraldehyde for 10 min, permeabilized with 0.1% Triton X-100 (Life Technologies) for 5 min, and stained with an anti-HbF APC antibody (Invitrogen) for 30 min. Cells were subsequently washed, acquired on an Accuri C6 flow cytometer (BD Biosciences), and analyzed using FlowJo software (v.10.8.1, BD Biosciences).…”

“…[2][3][4] These studies led to functional follow up that revealed how BCL11A acted as a key and direct repressor of HBG1/2 transcription. 5 Subsequently, additional insights have emerged from the study of rare loss-of-function mutations impacting BCL11A, [6][7][8] transcriptional regulatory elements necessary for erythroid expression of this factor, 9,10 upstream regulators of BCL11A expression, [11][12][13][14] and analysis of the mechanisms by which BCL11A alters transcription. [15][16][17] Suppression of BCL11A or its binding motif in the HBG1/2 promoters using genome editing or gene therapy approaches has emerged as a curative strategy for sickle cell disease and β-thalassemia.…”

Single Nucleotide Polymorphisms in XMN1-HBG2, HBS1L-MYB, and BCL11A and Their Relation to High Fetal Hemoglobin Levels That Alleviate Anemia

“…The remarkable hemoglobin shifting is influenced by the expression of the KLF1 (erythroid Kruppel-like factor) gene, which stimulates β-globin production and indirectly represses γ-globin expression [ 77 ]. However, downregulation of KLF1 in vivo has revealed a decrease in BCL11A expression in adult erythroid progenitors, which leads to activation of HbF production in anemic β-thalassemia, SCD, and HPFH patients [ 78 , 79 ].…”