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The human ζ-globin gene (HBZ) is transcribed in primitive erythroid cells only during the embryonic stages of development. Reactivation of this embryonic globin synthesis would likely alleviate symptoms both in α-thalassemia and sickle-cell disease. However, the molecular mechanisms controlling ζ-globin expression have remained largely undefined. Moreover, the pharmacologic agent capable of inducing ζ-globin production is currently unavailable. Here, we show that TRIAC, a bioactive thyroid hormone metabolite, significantly induced ζ-globin gene expression during zebrafish embryogenesis. The induction of ζ-globin expression by TRIAC was also observed in human K562 erythroleukemia cell line and primary erythroid cells. Thyroid hormone receptor α (THRA) deficiency abolished the ζ-globin-inducing effect of TRIAC. Furthermore, THRA could directly bind to the distal enhancer regulatory element to regulate ζ-globin expression. Our study provides the first evidence that TRIAC acts as a potent inducer of ζ-globin expression, which might serve as a new potential therapeutic option for patients with severe α-thalassemia or sickle-cell disease. Supplementary Information The online version contains supplementary material available at 10.1186/s13045-021-01108-z.

The α- and β-globin loci harbor developmentally expressed genes, which are silenced throughout post-natal life. Reactivation of these genes may offer therapeutic approaches for the hemoglobinopathies, the most common single gene disorders. Here, we address mechanisms regulating the embryonically expressed α-like globin, termed ζ-globin. We show that in embryonic erythroid cells, the ζ-gene lies within a ~65 kb sub-TAD (topologically associating domain) of open, acetylated chromatin and interacts with the α-globin super-enhancer. By contrast, in adult erythroid cells, the ζ-gene is packaged within a small (~10 kb) sub-domain of hypoacetylated, facultative heterochromatin within the acetylated sub-TAD and that it no longer interacts with its enhancers. The ζ-gene can be partially re-activated by acetylation and inhibition of histone de-acetylases. In addition to suggesting therapies for severe α-thalassemia, these findings illustrate the general principles by which reactivation of developmental genes may rescue abnormalities arising from mutations in their adult paralogues., Globin loci harbor genes that are expressed embryonically and silenced postnatally. Here the authors show that zeta-globin silencing depends upon selective hypoacetylation of its TAD subdomain, which blocks its interaction with the alpha-globin super-enhancer, and zeta-globin can be reactivated by acetylation.

BACKGROUND: Hemochromatosis gene (HFE)-associated hereditary hemochromatosis (HH) is characterized by downregulation of hepcidin synthesis, leading to increased intestinal iron absorption. OBJECTIVES: The objectives were to characterize and elucidate a possible association between gene expression profile, hepcidin levels, disease severity, and markers of inflammation in HFE-associated HH patients. METHODS: Thirty-nine HFE-associated HH patients were recruited and assigned to 2 groups according to genetic profile: C282Y homozygotes in 1 group and patients with H63D, as homozygote or in combination with C282Y, in the other group. Eleven healthy first-time blood donors were recruited as controls. Gene expression was characterized from peripheral blood cells, and inflammatory cytokines and hepcidin-25 isoform were quantified in serum. Biochemical disease characteristics were recorded. RESULTS: Elevated levels of interleukin 8 were observed in a significant higher proportion of patients than controls. In addition, compared to controls, gene expression of ζ-globin was significantly increased among C282Y homozygote patients, while gene expression of matrix metalloproteinase 8, and other neutrophil-secreted proteins, was significantly upregulated in patients with H63D. CONCLUSION: Different disease signatures may characterize HH patients according to their HFE genetic profile. Studies on larger populations, including analyses at protein level, are necessary to confirm these findings.

α0-Thalassemia is an inherited hematological disorder caused by the deletion of α-globin genes. The Southeast Asian deletion (--SEA) is the most common type of α0-thalassemia observed in Southeast Asian countries. Regarding WHO health policy, an effective α0-thalassemia screening strategy is needed to control new severe α-thalassemia cases. In this study, a monoclonal antibody panel was used to develop immunochromatographic (IC) strip tests for detecting the Hb Bart's and ζ-globin chain. Among 195 samples, all α0-thalassemia traits (78 α0-thalassemia (--SEA) and 4 α0-thalassemia (--THAI)) had low MCV or MCH values. The sensitivity, specificity, PPV and NPV of the IC strip tests for ζ-globin and Hb Bart's for screening α0-thalassemia (--SEA) within the low MCV or MCH samples were 100%, 65.2%, 90.7%, 100% and 96.2%, 47.8%, 86.6%, 78.6%, respectively. All 4 α0-thalassemia (--THAI) traits were negative for ζ-globin chains but positive for Hb Bart's using the IC strip tests. These results led to a α0-thalassemia screening being proposed in which blood samples are first evaluated by MCV, MCH and Hb typing. Samples with high MCV and MCH values are excluded for the presence of the α0-thalassemia gene. Samples with low MCV or MCH values are assayed using the developed IC strip tests, where only samples testing positive are further assayed for α0-thalassemia by PCR. Patients with Hb H, EA Bart's or EF Bart's diseases do not need to use this IC strip assay. Thus, in this study, a simple and cost effective α0-thalassemia point of care test was developed.

AimsThe presence of the ζ-globin chain is a good marker of (--SEA) α0-thalassaemia. We evaluated an immunochromatographic (IC) strip assay for ζ-globin in screening for (--SEA) α0-thalassaemia in a population with a high prevalence and heterogeneity of haemoglobinopathies.MethodsThe study was carried out on 300 screen positive blood samples of Thai individuals. The IC strip assay for the ζ-globin chain was performed on all samples. The results were interpreted with thalassaemia genotyping using standard haemoglobin and DNA analyses.ResultsSeveral thalassaemia genotypes were noted. Among the 300 subjects investigated, 79 had a positive IC strip assay for ζ-globin and (--SEA) α0-thalassaemia was identified in 40 of them. No (--SEA) α0-thalassaemia was detected in the remaining 39 samples with a positive IC strip test result or in the 221 samples with a negative IC strip test result. Further DNA analysis identified α+-thalassaemia in 25 of the 39 (--SEA) α0-thalassaemia negative samples. Using this IC strip assay in combination with a conventional screening protocol for (--SEA) α0-thalassaemia could provide sensitivity and specificity of 100% and 90.4%, respectively.ConclusionsIC strip assay for ζ-globin is simple, rapid and does not require sophisticated equipment. Use of this test in addition to the existing screening protocol could detect potential (--SEA) α0-thalassaemia leading to a significant reduction in the workload of DNA analysis. This could be used in areas where haemoglobinopathies are prevalent and heterogeneous but molecular testing is not available.

A reliable cell line capable of robust in vitro erythroid differentiation would be useful to investigate red blood cell (RBC) biology and genetic strategies for RBC diseases. K562 cells are widely utilized for erythroid differentiation; however, current differentiation methods are insufficient to analyze globin proteins. In this study, we sought to improve erythroid differentiation from K562 cells to enable protein-level globin analysis. K562 cells were exposed to a variety of reagents, including hemin, rapamycin, imatinib, and/or decitabine (known erythroid inducers), and cultured in a basic culture medium or erythropoietin-based differentiation medium. All single reagents induced observable erythroid differentiation with higher glycophorin A (GPA) expression but were insufficient to produce detectable globin proteins. We then evaluated various combinations of these reagents and developed a method incorporating imatinib preexposure and an erythropoietin-based differentiation culture containing both rapamycin and decitabine capable of efficient erythroid differentiation, high-level GPA expression (>90%), and high-level globin production at protein levels detectable by hemoglobin electrophoresis and high performance liquid chromatography. In addition, β-globin gene transfer resulted in detectable adult hemoglobin. In summary, we developed an in vitro K562 erythroid differentiation model with high-level globin production. This model provides a practical evaluation tool for hemoglobin production in human erythroid cells.