Drosophila O-GlcNAcase Deletion Globally Perturbs Chromatin O-GlcNAcylation
Drosophila O-GlcNAcase Deletion Globally Perturbs Chromatin O-GlcNAcylation
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DOI：
10.1074/jbc.m115.704783
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发表时间：
2016-05-06
影响因子：
4.8
通讯作者：
Hanover, John A.
中科院分区：
文献类型：
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作者：
Akan, Ilhan;Love, Dona C.;Hanover, John A.
果蝇发育过程中的基因表达受到Polycomb（Pc）、Trithorax（Trx）和Compass染色质修饰复合物的调控。O-GlcNAc转移酶（OGT/SXC）是Pc抑制所必需的，表明蛋白质的O-GlcNAc化在调节发育中起关键作用。OGT将O-GlcNAc转移到关键转录调节因子的内在无序结构域中的丝氨酸和苏氨酸残基上; O-GlcNAc酶（OGA）去除了修饰。为了查明受O-GlcNAc水平调控的基因组区域，我们在S2细胞中OGT或OGA RNAi敲低后进行了ChIP芯片和微阵列分析。在OGA RNAi后，我们观察到大多数O-GlcNAc占据区域的基因组范围内的强度增加，包括与细胞周期，泛素和类固醇反应相关的基因。相比之下，O-GlcNAc水平对多梳抑制位点（如Hox和NK同源框基因簇）的OGA RNAi显著不敏感。微阵列分析表明，改变O-GlcNAc循环扰乱与形态发生和细胞周期调控相关的基因的表达。然后，我们在果蝇中产生了一个可行的无效等位基因oga（oga（del.1）），允许观察多线染色体上改变的O-GlcNAc循环。我们发现，三胸（TRX），缺乏小或同源异型盘1（ASH 1），和指南针成员SET 1组蛋白甲基转移酶O-GlcNAc修饰的oga（del.1）突变体。oga（del.1）突变体显示了一组不同的细胞周期相关基因的表达改变。我们的研究结果表明，在果蝇中的OGA的损失全球影响的表观遗传机制，允许O-GlcNAc积累的RNA聚合酶II和许多染色质因子，包括TRX，ASH 1和SET 1。
Gene expression during Drosophila development is subject to regulation by the Polycomb (Pc), Trithorax (Trx), and Compass chromatin modifier complexes. O-GlcNAc transferase (OGT/SXC) is essential for Pc repression suggesting that the O-GlcNAcylation of proteins plays a key role in regulating development. OGT transfers O-GlcNAc onto serine and threonine residues in intrinsically disordered domains of key transcriptional regulators; O-GlcNAcase (OGA) removes the modification. To pinpoint genomic regions that are regulated by O-GlcNAc levels, we performed ChIP-chip and microarray analysis after OGT or OGA RNAi knockdown in S2 cells. After OGA RNAi, we observed a genome-wide increase in the intensity of most O-GlcNAc-occupied regions including genes linked to cell cycle, ubiquitin, and steroid response. In contrast, O-GlcNAc levels were strikingly insensitive to OGA RNAi at sites of polycomb repression such as the Hox and NK homeobox gene clusters. Microarray analysis suggested that altered O-GlcNAc cycling perturbed the expression of genes associated with morphogenesis and cell cycle regulation. We then produced a viable null allele of oga (oga(del.1)) in Drosophila allowing visualization of altered O-GlcNAc cycling on polytene chromosomes. We found that trithorax (TRX), absent small or homeotic discs 1 (ASH1), and Compass member SET1 histone methyltransferases were O-GlcNAc-modified in oga(del.1) mutants. The oga(del.1) mutants displayed altered expression of a distinct set of cell cycle-related genes. Our results show that the loss of OGA in Drosophila globally impacts the epigenetic machinery allowing O-GlcNAc accumulation on RNA polymerase II and numerous chromatin factors including TRX, ASH1, and SET1.