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 Gγ‐ and β‐globin genes are replaced by DsRed and eGFP fluorescent reporters, respectively. Following RNA interference (RNAi)‐mediated 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 RNAi 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.—Roosjen, M., McColl, B., Kao, B., Gearing, L. J., Blewitt, M. E., Vadolas, J. Transcriptional regulators Myb and BCL11A interplay with DNA methyltransferase 1 in developmental silencing of embryonic and fetal β‐like globin genes. FASEB J. 28, 28–1610 (1620). www.fasebj.org