Distributive O-GlcNAcylation on the Highly Repetitive C-Terminal Domain of RNA Polymerase II.
Distributive O-GlcNAcylation on the Highly Repetitive C-Terminal Domain of RNA Polymerase II.
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DOI：
10.1021/acs.biochem.5b01280
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发表时间：
2016-02-23
期刊：
影响因子：
2.9
通讯作者：
Jiang J
中科院分区：
文献类型：
--
作者：
Lu L;Fan D;Hu CW;Worth M;Ma ZX;Jiang J
O-GlcNAcylation是一种营养响应性糖基化，在转录调控中起关键作用。人RNA聚合酶II（Pol II）在其独特的C-末端结构域（CTD）上被O-连接的N-乙酰葡糖胺（O-GlcNAc）广泛修饰，该结构域由52个七肽重复组成。了解糖基化Pol II功能的一种方法是确定CTD上动态O-GlcNAc酰化的机制。在这里，我们发现Pol II CTD可以在体外和细胞中被广泛地O-GlcNAc酰化。有效的糖基化需要至少20个七肽重复的CTD和超过一半的O-GlcNAc转移酶（OGT）N末端结构域。在饱和糖供体的条件下，我们监测到超过20个残基的O-GlcNAc连接到全长CTD。令人惊讶的是，周期性CTD上的糖基化遵循分布机制，导致高度异质的糖型。我们的数据表明，初始O-GlcNAc酰化可以发生在CTD的近端或远端区域，随后的糖基化类似地发生在整个CTD上，具有不均匀的分布。此外，从糖基化CTD中去除O-GlcNAc也是分布性的，并且与O-GlcNAc化水平无关。我们的研究结果表明，O-GlcNAc循环酶可以采用类似的机制与其他蛋白质底物在多个位点上反应。Pol II上的分布性O-GlcNAc化提供了响应波动的细胞条件的另一种转录调节机制。
O-GlcNAcylation is a nutrient-responsive glycosylation that plays a pivotal role in transcriptional regulation. Human RNA polymerase II (Pol II) is extensively modified by O-linked N-acetylglucosamine (O-GlcNAc) on its unique C-terminal domain (CTD), which consists of 52 heptad repeats. One approach to understanding the function of glycosylated Pol II is to determine the mechanism of dynamic O-GlcNAcylation on the CTD. Here, we discovered that the Pol II CTD can be extensively O-GlcNAcylated in vitro and in cells. Efficient glycosylation requires a minimum of 20 heptad repeats of the CTD and more than half of the N-terminal domain of O-GlcNAc transferase (OGT). Under conditions of saturated sugar donor, we monitored the attachment of more than 20 residues of O-GlcNAc to the full-length CTD. Surprisingly, glycosylation on the periodic CTD follows a distributive mechanism, resulting in highly heterogeneous glycoforms. Our data suggest that initial O-GlcNAcylation can take place either on the proximal or on the distal region of the CTD, and subsequent glycosylation occurs similarly over the entire CTD with nonuniform distributions. Moreover, removal of O-GlcNAc from glycosylated CTD is also distributive and is independent of O-GlcNAcylation level. Our results suggest that O-GlcNAc cycling enzymes can employ a similar mechanism to react with other protein substrates on multiple sites. Distributive O-GlcNAcylation on Pol II provides another regulatory mechanism of transcription in response to fluctuating cellular conditions.
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DOI：
10.1083/jcb.201501101
发表时间：
2015-03-30
期刊：
The Journal of cell biology
影响因子：
--
作者：
Bond MR;Hanover JA
通讯作者：
Hanover JA
影响因子：
3.8
作者：
Ma J;Hart GW
通讯作者：
Hart GW
影响因子：
4.8
作者：
Ranuncolo, Stella M.;Ghosh, Salil;Lewis, Brian A.
通讯作者：
Lewis, Brian A.
影响因子：
56.9
作者：
Cramer, P;Bushnell, DA;Kornberg, RD
通讯作者：
Kornberg, RD
影响因子：
14.8
作者：
Rexach, Jessica E.;Clark, Peter M.;Mason, Daniel E.;Neve, Rachael L.;Peters, Eric C.;Hsieh-Wilson, Linda C.
通讯作者：
Hsieh-Wilson, Linda C.