Fine Tuning of Globin Gene Expression by DNA Methylation

Expression patterns in the globin gene cluster are subject to developmental regulation in vivo. While the γ(A) and γ(G) genes are expressed in fetal liver, both are silenced in adult erythrocytes. In order to decipher the role of DNA methylation in this process, we generated a YAC transgenic mouse system that allowed us to control γ(A) methylation during development. DNA methylation causes a 20-fold repression of γ(A) both in non-erythroid and adult erythroid cells. In erythroid cells this modification works as a dominant mechanism to repress γ gene expression, probably through changes in histone acetylation that prevent the binding of erythroid transcription factors to the promoter. These studies demonstrate that DNA methylation serves as an elegant in vivo fine-tuning device for selecting appropriate genes in the globin locus. In addition, our findings provide a mechanism for understanding the high levels of γ-globin transcription seen in patients with Hereditary Persistence of Fetal Hemoglobin, and help explain why 5azaC and butyrate compounds stimulate γ-globin expression in patients with β-hemoglobinopathies

Persistent Fetal γ-Globin Expression in Adult Transgenic Mice following Deletion of Two Silencer Elements Located 3′ to the Human Aγ-Globin Gene

Natural deletions of the human γ-globin gene cluster lead to specific syndromes characterized by increased production of fetal hemoglobin in adult life and provide a useful model to delineate novel cis-acting elements involved in the developmental control of hemoglobin switching. A hypothesis accounting for these phenotypic features assumes that silencers located within the Aγ-to δ-gene region are deleted in hereditary persistence of fetal hemoglobin (HPFH) and δβ-thalassemias, leading to failure of switching. In the present study, we sought to clarify the in vivo role of two elements, termed Enh and F, located 3′ to the Aγ-globin, in silencing the fetal genes. To this end, we generated three transgenic lines using cosmid constructs containing the full length of the globin locus control region (LCR) linked to the 3.3-kb Aγ-gene lacking both the Enh and F elements. The Enh/F deletion resulted in high levels of Aγ-globin gene expression in adult mice in all single copy lines, whereas, the LCR-Aγ single copy lines which retain the Enh and F elements exhibited complete normal switching of the fetal Aγ-gene. Our study documents directly for the first time the in vivo role of these two gene-proximal negative regulatory elements in silencing the fetal globin gene in the perinatal period, and thus these data may permit their eventual exploitation in therapeutic approaches for thalassemias