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O-Linked N-Acetylglucosamine (O-GlcNAc) Transferase and O-GlcNAcase Interact with Mi2β Protein at the Aγ-Globin Promoter

2016; Elsevier BV; Volume: 291; Issue: 30 Linguagem: Inglês

10.1074/jbc.m116.721928

1083-351X

Zhen Zhang, Flavia C Costa, Ee Phie Tan, Nathan Bushue, Luciano DiTacchio, Catherine E. Costello, Mark E. McComb, Stephen A. Whelan, Kenneth R. Peterson, Chad Slawson,

Carbohydrate Chemistry and Synthesis

One mode of γ-globin gene silencing involves a GATA-1·FOG-1·Mi2β repressor complex that binds to the −566 GATA site relative to the Aγ-globin gene cap site. However, the mechanism of how this repressor complex is assembled at the −566 GATA site is unknown. In this study, we demonstrate that the O-linked N-acetylglucosamine (O-GlcNAc) processing enzymes, O-GlcNAc-transferase (OGT) and O-GlcNAcase (OGA), interact with the Aγ-globin promoter at the −566 GATA repressor site; however, mutation of the GATA site to GAGA significantly reduces OGT and OGA promoter interactions in β-globin locus yeast artificial chromosome (β-YAC) bone marrow cells. When WT β-YAC bone marrow cells are treated with the OGA inhibitor Thiamet-G, the occupancy of OGT, OGA, and Mi2β at the Aγ-globin promoter is increased. In addition, OGT and Mi2β recruitment is increased at the Aγ-globin promoter when γ-globin becomes repressed in postconception day E18 human β-YAC transgenic mouse fetal liver. Furthermore, we show that Mi2β is modified with O-GlcNAc, and both OGT and OGA interact with Mi2β, GATA-1, and FOG-1. Taken together, our data suggest that O-GlcNAcylation is a novel mechanism of γ-globin gene regulation mediated by modulating the assembly of the GATA-1·FOG-1·Mi2β repressor complex at the −566 GATA motif within the promoter. One mode of γ-globin gene silencing involves a GATA-1·FOG-1·Mi2β repressor complex that binds to the −566 GATA site relative to the Aγ-globin gene cap site. However, the mechanism of how this repressor complex is assembled at the −566 GATA site is unknown. In this study, we demonstrate that the O-linked N-acetylglucosamine (O-GlcNAc) processing enzymes, O-GlcNAc-transferase (OGT) and O-GlcNAcase (OGA), interact with the Aγ-globin promoter at the −566 GATA repressor site; however, mutation of the GATA site to GAGA significantly reduces OGT and OGA promoter interactions in β-globin locus yeast artificial chromosome (β-YAC) bone marrow cells. When WT β-YAC bone marrow cells are treated with the OGA inhibitor Thiamet-G, the occupancy of OGT, OGA, and Mi2β at the Aγ-globin promoter is increased. In addition, OGT and Mi2β recruitment is increased at the Aγ-globin promoter when γ-globin becomes repressed in postconception day E18 human β-YAC transgenic mouse fetal liver. Furthermore, we show that Mi2β is modified with O-GlcNAc, and both OGT and OGA interact with Mi2β, GATA-1, and FOG-1. Taken together, our data suggest that O-GlcNAcylation is a novel mechanism of γ-globin gene regulation mediated by modulating the assembly of the GATA-1·FOG-1·Mi2β repressor complex at the −566 GATA motif within the promoter.