Transcriptional regulators Myb and BCL11A interplay with DNA methyltransferase 1 in developmental silencing of embryonic and fetal β-like globin genes.

The clinical symptoms of hemoglobin disorders such as β-thalassemia and sickle cell anemia are significantly ameliorated by the persistent expression of γ-globin after birth. This knowledge has driven the discovery of important regulators that silence γ-globin postnatally. Improved understanding of the γ- to β-globin switching mechanism holds the key to devising targeted therapies for β-hemoglobinopathies. To further investigate this mechanism, we used the murine erythroleukemic (MEL) cell line containing an intact 183-kb human β-globin locus, in which the (G)γ- and β-globin genes are replaced by DsRed and eGFP fluorescent reporters, respectively. Following RNA interference knockdown of two key transcriptional regulators, Myb and BCL11A, we observed a derepression of γ-globin, measured by DsRed fluorescence and qRT-PCR (P<0.001). Interestingly, double knockdown of Myb and DNA methyltransferase 1 (DNMT1) resulted in a robust induction of ε-globin, (up to 20% of total β-like globin species) compared to single knockdowns (P<0.001). Conversely, double knockdowns of BCL11A and DNMT1 enhanced γ-globin expression (up to 90% of total β-like globin species) compared to single knockdowns (P<0.001). Moreover, following treatment, expression of human β-like globin genes mirrored the expression levels of their endogenous murine counterparts. These results demonstrate that Myb and BCL11A cooperate with DNMT1 to achieve developmental repression of embryonic and fetal β-like globin genes in the adult erythroid environment.

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