5-azacytidine treatment and TaPBF-D over-expression increases glutenin accumulation within the wheat grain by hypomethylating the Glu-1 promoters
5-azacytidine treatment and TaPBF-D over-expression increases glutenin accumulation within the wheat grain by hypomethylating the Glu-1 promoters
复制标题
5-氮杂胞苷处理和 TaPBF-D 过表达通过 Glu-1 启动子的低甲基化增加了小麦籽粒内麦谷蛋白的积累
DOI：
10.1007/s00122-017-3032-z
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发表时间：
2017
期刊：
影响因子：
--
通讯作者：
夏光敏
中科院分区：
文献类型：
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作者：
朱建堂;房琳琳;于佳锜;赵莹;陈凡国;夏光敏
关键信息5-azaC处理和TaPBF-Dover表达降低了3个Glu-1基因启动子的C-甲基化状态，并有助于增强Glu-1基因的表达。摘要小麦麦谷蛋白对面团弹性有很强的影响，但其编码基因的调控尚未确定。经5-氮胞苷(5-azaC)处理后，发育期和成熟期籽粒的重量和谷蛋白含量均显著增加。Glu-1基因(编码高分子量谷蛋白亚基)以及编码去甲基酶和醇溶蛋白合成相关转录因子的转录本的丰度高于未处理植株的籽粒。这些谷物还含有增加的醇溶蛋白盒结合因子(PBF)蛋白。亚硫酸氢盐测序表明，Glu-1启动子在发育籽粒中的C-甲基化程度低于旗叶，而在5-azaC处理植株的发育籽粒中，C-甲基化水平低于未处理植株的相应籽粒。与野生型植株相比，TaPBF D基因组同源基因的3个独立过表达基因所形成的籽粒中Glu-1转录本丰度和谷蛋白含量均较高。在花后10天进行评估时，在TaPBF-Dover表达载体设置的发育中的籽粒中，Glu-1启动子的甲基化水平低于野生型对照。电泳迁移率改变分析表明，PBF-D能够在体外与Glu-1By8和-1Dx2的P盒结合，而芯片定量PCR分析表明，Glu-1By8和-1Dx2启动子中较低水平的C-甲基化改善了TaPBF的结合。我们认为启动子DNA C-甲基化是Glu-1转录的关键决定因素。
Key message5-azaC treatment andTaPBF-Dover-expression decrease C-methylation status of threeGlu-1gene promoters, and aid in enhancing the expression of theGlu-1genes.AbstractThe wheat glutenins exert a strong influence over dough elasticity, but the regulation of their encoding genes has not been firmly established. Following treatment with 5-azacytidine (5-azaC), both the weight and glutenin content of the developing and mature grains were significantly increased. The abundance of transcript produced by theGlu-1genes (encoding high-molecular-weight glutenin subunits), as well as those encoding demethylases and transcriptional factors associated with prolamin synthesis was higher than in grain of non-treated plants. These grains also contained an enhanced content of the prolamin box binding factor (PBF) protein. Bisulfite sequencing indicated that theGlu-1promoters were less strongly C-methylated in the developing grain than in the flag leaf, while in the developing grain of 5-azaC treated plants, the C-methylation level was lower than in equivalent grains of non-treated plants. BothGlu-1transcript abundance and glutenin content were higher in the grain set by three independent over-expressors of the D genome homoeolog ofTaPBFthan in the grain set by wild type plants. When assessed 10 days after flowering, theGlu-1promoters’ methylation level was lower in the developing grains set by theTaPBF-Dover-expressor than in the wild type control. An electrophoretic mobility shift assay showed that PBF-D was able to bind in vitro to the P-box ofGlu-1By8and -1Dx2, while a ChIP-qPCR analysis revealed that a lower level of C-methylation in theGlu-1By8and -1Dx2promoters improved the TaPBF binding. We suggest that promoter DNA C-methylation is a key determinant ofGlu-1transcription.