Introducing the −113A>G HPFH mutation into erythroid cells elevates fetal globin levels.
The −113A>G mutation creates a de novo site for the activator GATA1 but does not disrupt binding of fetal globin repressor BCL11A.
β-hemoglobinopathies, such as sickle cell disease and β-thalassemia, result from mutations in the adult β-globin gene. Reactivating the developmentally silenced fetal γ-globin gene elevates fetal hemoglobin levels and ameliorates symptoms of β-hemoglobinopathies. The continued expression of fetal γ-globin into adulthood occurs naturally in a genetic condition termed hereditary persistence of fetal hemoglobin (HPFH). Point mutations in the fetal γ-globin proximal promoter can cause HPFH. The −113A>G HPFH mutation falls within the −115 cluster of HPFH mutations, a binding site for the fetal globin repressor BCL11A. We demonstrate that the −113A>G HPFH mutation, unlike other mutations in the cluster, does not disrupt BCL11A binding but rather creates a de novo binding site for the transcriptional activator GATA1. Introduction of the −113A>G HPFH mutation into erythroid cells using the clustered regularly interspaced short palindromic repeats (CRISPR)–CRISPR-associated protein 9 (Cas9) system increases GATA1 binding and elevates fetal globin levels. These results reveal the mechanism by which the −113A>G HPFH mutation elevates fetal globin and demonstrate the sensitivity of the fetal globin promoter to point mutations that often disrupt repressor binding sites but here create a de novo site for an erythroid activator.