Ye Juanying, Zhang Zaibao, You Chenjiang, Zhang Xumin, Lu Jianan, Ma Hong
State Key Laboratory of Genetic Engineering and Collaborative Innovation Center of Genetics and Development, Institute of Plant Biology, School of Life Sciences, Fudan University, Shanghai 200433, China.
State Key Laboratory of Genetic Engineering and Collaborative Innovation Center of Genetics and Development, Institute of Plant Biology, School of Life Sciences, Fudan University, Shanghai 200433, China
J Exp Bot. 2016 Sep;67(17):4993-5008. doi: 10.1093/jxb/erw293. Epub 2016 Aug 16.
As the male reproductive organ of flowering plants, the stamen consists of the anther and filament. Previous studies on stamen development mainly focused on single gene functions by genetic methods or gene expression changes using comparative transcriptomic approaches, especially in model plants such as Arabidopsis thaliana However, studies on Arabidopsis anther protein expression and post-translational modifications are still lacking. Here we report proteomic and phosphoproteomic studies on developing Arabidopsis anthers at stages 4-7 and 8-12. We identified 3908 high-confidence phosphorylation sites corresponding to 1637 phosphoproteins. Among the 1637 phosphoproteins, 493 were newly identified, with 952 phosphorylation sites. Phosphopeptide enrichment prior to LC-MS analysis facilitated the identification of low-abundance proteins and regulatory proteins, thereby increasing the coverage of proteomic analysis, and facilitated the analysis of more regulatory proteins. Thirty-nine serine and six threonine phosphorylation motifs were uncovered from the anther phosphoproteome and further analysis supports that phosphorylation of casein kinase II, mitogen-activated protein kinases, and 14-3-3 proteins is a key regulatory mechanism in anther development. Phosphorylated residues were preferentially located in variable protein regions among family members, but they were they were conserved across angiosperms in general. Moreover, phosphorylation might reduce activity of reactive oxygen species scavenging enzymes and hamper brassinosteroid signaling in early anther development. Most of the novel phosphoproteins showed tissue-specific expression in the anther according to previous microarray data. This study provides a community resource with information on the abundance and phosphorylation status of thousands of proteins in developing anthers, contributing to understanding post-translational regulatory mechanisms during anther development.
作为开花植物的雄性生殖器官，雄蕊由花药和花丝组成。以往对雄蕊发育的研究主要集中在通过遗传方法研究单个基因的功能，或使用比较转录组学方法研究基因表达变化，特别是在拟南芥等模式植物中。然而，关于拟南芥花药蛋白质表达和翻译后修饰的研究仍然缺乏。在这里，我们报告了对处于4-7期和8-12期的拟南芥发育花药的蛋白质组学和磷酸化蛋白质组学研究。我们鉴定出了与1637个磷酸化蛋白质相对应的3908个高可信度磷酸化位点。在这1637个磷酸化蛋白质中，新鉴定出493个，具有952个磷酸化位点。液相色谱-质谱分析前的磷酸肽富集有助于鉴定低丰度蛋白质和调节蛋白，从而增加蛋白质组分析的覆盖范围，并有助于分析更多的调节蛋白。从花药磷酸化蛋白质组中发现了39个丝氨酸和6个苏氨酸磷酸化基序，进一步分析支持酪蛋白激酶II、丝裂原活化蛋白激酶和14-3-3蛋白的磷酸化是花药发育中的关键调节机制。磷酸化残基优先位于家族成员间可变的蛋白质区域，但总体上在被子植物中是保守的。此外，磷酸化可能会降低活性氧清除酶的活性，并在花药发育早期阻碍油菜素内酯信号传导。根据之前的微阵列数据，大多数新的磷酸化蛋白质在花药中表现出组织特异性表达。这项研究提供了一个社区资源，包含发育花药中数千种蛋白质的丰度和磷酸化状态信息，有助于理解花药发育过程中的翻译后调控机制。
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