藤木 亮次

The post-translational modifications of histone tails generate a 'histone code' that defines local and global chromatin states(1). The resultant regulation of gene function is thought to govern cell fate, proliferation and differentiation(2). Reversible histone modifications such as methylation are under mutual controls to organize chromosomal events(3,4). Among the histone modifications, methylation of specific lysine and arginine residues seems to be critical for chromatin configuration and control of gene expression(5). Methylation of histone H3 lysine 4 (H3K4) changes chromatin into a transcriptionally active state(6). Reversible modification of proteins by beta-N-acetylglucosamine (O-GlcNAc) in response to serum glucose levels regulates diverse cellular processes(7,8,9). However, the epigenetic impact of protein GlcNAcylation is unknown. Here we report that nuclear GlcNAcylation of a histone lysine methyltransferase (HKMT), MLL5, by O-GlcNAc transferase facilitates retinoic-acid-induced granulopoiesis in human HL60 promyelocytes through methylation of H3K4. MLL5 is biochemically identified in a GlcNAcylation-dependent multi-subunit complex associating with nuclear retinoic acid receptor RAR alpha (also known as RARA), serving as a mono-and di-methyl transferase to H3K4. GlcNAcylation at Thr 440 in the MLL5 SET domain evokes its H3K4 HKMT activity and co-activates RARa in target gene promoters. Increased nuclear GlcNAcylation by means of O-GlcNAc transferase potentiates retinoic-acid-induced HL60 granulopoiesis and restores the retinoic acid response in the retinoic-acid-resistant HL60-R2 cell line. Thus, nuclear MLL5 GlcNAcylation triggers cell lineage determination of HL60 through activation of its HKMT activity.

We have previously shown that the novel ATP-dependent chromatin remodeling complex WINAC is required for the ligand-bound Vitamin D receptor (VDR)-mediated transrepression of the 25(OH)D-3 1 alpha-hydroxylase [1 alpha(OH)ase] gene. However, the molecular basis for VDR promoter association, which does not involve its binding to specific DNA sequences, remains unclear. To address this issue, we investigated the function of WSTF in terms of the association between WINAC and chromatin for ligand-induced transrepression by VDR. Results of in vitro experiments using chromatin templates showed that the association of unliganded VDR with the promoter required physical interactions between WSTF and both VDR and acetylated histones prior to VDR association with chromatin. The acetylated histone-interacting region of WSTF was mapped to the bromodomain, and a WSTF mutant lacking the bromodomain served as a dominant-negative mutant in terms of ligand-induced transrepression of the lot(OH)ase gene. Thus, our findings indicate that WINAC associates with chromatin through a physical interaction between the WSTF bromodomain and acetylated histones, that appears to be indispensable for VDR/promoter association for ligand-induced transrepression of 1 alpha(OH)ase gene expression. (c) 2006 Elsevier Ltd. All rights reserved.

CYP27B1 is a critical enzyme of Vitamin D biosynthesis that hydroxylates 25(OH)D-3 at the final step of the biosynthetic pathway. The CYP27B1 gene is expressed primarily in kidney and negatively controlled by Vitamin D receptor. We have characterized the negative vitamin D response element and its binding protein, a bHLH transcription factor. This factor directly binds to the 1 alpha nVDRE and activates transcription, but its transcriptional activity is suppressed by the ligand-activated Vitamin D receptor through recruitment of histone deacetylase. We have shown that histone deacetylation is a critical step for chromatin structure remodeling in suppression of the CYP27B1 gene. We have further demonstrated that, in addition to histone acetylation, this transrepression by VDR requires DNA methylation in the CYP27B1 gene promoter. Thus, transcriptional regulation of the CYP27B1 gene appears to be mediated by dual epigenetic modifications. (c) 2006 Elsevier Ireland Ltd. All rights reserved.