NR2F1 : A Novel Candidate for Gamma Globin Activation

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The reactivation of human gamma globin holds tremendous promise as a potential treatment for hemoglobinopathies, including sickle cell disease and beta-thalassemia. Despite identifying various repressors of fetal globin genes in adult erythroid cells over the past decade, evidence concerning stage-specific factors that activate human fetal globin prior to the switch to adult beta-globin remains inadequately understood. In our study, we constructed an erythroid-specific gene regulatory network (GCN) by integrating multiple RNA-seq datasets from erythroid cells at different stages of differentiation and development. Our analysis unveiled both known HbF activators, like HIC2, and novel candidates. Among the novel contenders, NR2F1, a member of the steroid/thyroid hormone receptor superfamily, emerged as a prominent candidate. To validate our predictions, we overexpressed NR2F1 in Hudep2 cells and observed more than a four-fold increase (28% vs. 6%) in the proportion of F cells and a three-fold rise in HbF levels as measured by HPLC. In addition, the overexpression of NR2F1 did not appear to impact beta-globin expression or cell differentiation. Intriguingly, we observed a significant reduction in the protein level of BCL11A, a master regulator of HbF, despite no major change in the total RNA level of BCL11A. This observation suggests that NR2F1 plays a role in the post-transcriptional modification of BCL11A. Additionally, NR2F1 overexpression led to an upregulation of Lin28B expression in Hudep2 cells. Lin28B is known to directly bind BCL11A mRNA and prevent its effective translation without disturbing BCL11A transcription. Consequently, the observed upregulation of Lin28B in NR2F1-overexpressing cells implies a potential NR2F1-Lin28B-BCL11A axis involved in controlling human hemoglobin switching. In conclusion, our findings propose NR2F1 as a novel activator of gamma globin, representing an attractive target for gamma globin reactivation. Additional investigation is required to explore the molecular mechanisms through which NR2F1 regulates gamma globin expression and its role in the NR2F1-Lin28B-BCL11A axis.