O-Linked N-acetylglucosamine transferase 1 regulates global histone H4 acetylation via stabilization of the non-specific lethal protein NSL3 [Enzymology]

The human MOF-containing histone acetyltransferase (HAT) NSL (non-specific lethal) complex comprises 9 subunits including the O-linked N-acetylglucosamine (O-GlcNAc) transferase (isoform 1) (OGT1). However, whether O-GlcNAc transferase activity of OGT1 controls HAT activity of the NSL complex and whether OGT1 physically interacts with the other NSL complex subunits remains unclear. Here, we demonstrate that OGT1 regulates the activity of the NSL complex by mainly acetylating histone H4K16, K5, and K8 via O-GlcNAcylation and stabilization of the NSL complex subunit NSL3. Knocking down or overexpressing OGT1 in human cells remarkably affected the global acetylation of histone H4 residues K16, K5, and K8. Because OGT1 is a subunit of the NSL complex, we also investigated the function of OGT1 in this complex. Co-transfection/co-immunoprecipitation experiments combined with in vitro O-GlcNAc transferase assays confirmed that OGT1 specifically binds to and O-GlcNAcylates NSL3. In addition, WGA-affinity purification clarified the occurrence of O-GlcNAc modification on NSL3 in cells. Moreover, O-GlcNAcylation of NSL3 by wild type OGT1 (OGT1wt) stabilized NSL3. This stabilization was lost after co-transfection of NSL3 with an OGT1 mutant, OGT1C964A that lacks O-GlcNAc transferase activity. Furthermore, stabilization of NSL3 by OGT1wt significantly increased the global acetylation levels of H4K5, K8, and K16 in cells. These results suggest that OGT1 regulates the activity of the NSL complex by stabilizing NSL3.

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