The molecular mechanisms that control the switching of globin gene expression during development are fundamental to any efforts to design targeted therapy for the major hemoglobin disorders--sickle cell anemia and ?-thalassemia. Despite efforts of many investigators for more than two decades, the nuclear factors that regulate globin switches have remained largely elusive. Over the past two years genome-wide association studies examining the correlation of single-nucleotide polymorphisms with fetal hemoglobin (HbF) levels in populations have led to the identification of a candidate regulator of HbF expression, a gene product encoded by the BCL11A locus on chromosome 2. Previously we provided preliminary evidence in support of the hypothesis that BCL11A serves as a repressor of ?-globin expression. Down-regulation of BCL11A in adult CD34-derived erythroid precursors was associated with marked induction of ?-globin and HbF in culture. Our further studies have extended these findings by showing that BCL11A is the factor that restricts mouse embryonic ?-like globin expression to the primitive erythroid lineage and is required for the silencing of ?-globin expression in mice harboring a transgene with the entire human ?-globin locus. Thus, the aims of the current application focus specifically on how BCL11A functions to regulate these globin switches, as an improved understanding will provide critical insights into potential strategies to target down-regulation or inhibition of BCL11A expression or function to reactivate ?-globin expression in patients with the major hemoglobin disorders.
The aims of this project address the mechanisms by which BCL11A acts in erythroid cells.
Aim 1 addresses the structure and potential function of a unique isoform of BCL11A that is present in human embryonic and FL erythroid cells, but absent in mouse. This isoform may account in part for differences in switching of globins between mouse and man.
Aim 2 will define the functional domains within the BCL11A protein required for its function in ?-globin repression.
Aim 3 will determine to what extent human ?-globin expression can be reactivated in adult erythroid cells within the context of mouse ?-YAC transgenics.
Aim 4 will identify critical cis-regulatory elements within the human ?-locus required for proper ?-silencing by BCL11A.
Aim 5 will interrogate potential relationships between BCL11A and Sox6 in the control of globin switching. In aggregate, accomplishment of these aims will deepen a mechanistic understanding of globin switching and provide the necessary foundation for the design of targeted therapy for HbF reactivation.
(public) Red blood cells are products of what is called the erythroid lineage. Diseases of the erythroid lineage include the major disorders of hemoglobin (the oxygen carrying protein red blood cells) known as sickle cell anemia and the thalassemia syndromes. Our work is directed toward understanding how the red blood cell is programmed to express different types of hemoglobin at different times of development. Reactivation of a fetal form of hemoglobin (HbF) can ameliorate the consequences of sickle cell anemia and thalassemia. The work proposed in this application addresses directly how HbF is silenced normally in development by a specific protein known as BCL11A. This protein serves as the critical molecular switch in this process.
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|Canver, Matthew C; Lessard, Samuel; Pinello, Luca et al. (2017) Variant-aware saturating mutagenesis using multiple Cas9 nucleases identifies regulatory elements at trait-associated loci. Nat Genet 49:625-634|
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|Hoban, Megan D; Orkin, Stuart H; Bauer, Daniel E (2016) Genetic treatment of a molecular disorder: gene therapy approaches to sickle cell disease. Blood 127:839-48|
|Smith, Elenoe C; Orkin, Stuart H (2016) Hemoglobin genetics: recent contributions of GWAS and gene editing. Hum Mol Genet 25:R99-R105|
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|Luc, Sidinh; Huang, Jialiang; McEldoon, Jennifer L et al. (2016) Bcl11a Deficiency Leads to Hematopoietic Stem Cell Defects with an Aging-like Phenotype. Cell Rep 16:3181-3194|
|Orkin, Stuart H (2016) Recent advances in globin research using genome-wide association studies and gene editing. Ann N Y Acad Sci 1368:5-10|
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