Proteomic Approach to Identify Nuclear Proteins in Wheat Grain

The nuclear proteome of the grain of the two cultivated wheat species Triticum aestivum (hexaploid wheat; genomes A, B, and D) and T. monococcum (diploid wheat; genome A) was analyzed in two early stages of development using shotgun-based proteomics. A procedure was optimized to purify nuclei, and an improved protein sample preparation was developed to efficiently remove nonprotein substances (starch and nucleic acids). A total of 797 proteins corresponding to 528 unique proteins were identified, 36% of which were classified in functional groups related to DNA and RNA metabolism. A large number (107 proteins) of unknown functions and hypothetical proteins were also found. Some identified proteins may be multifunctional and may present multiple localizations. On the basis of the MS/MS analysis, 368 proteins were present in the two species, and in two stages of development, some qualitative differences between species and stages of development were also found. All of these data illustrate the dynamic function of the grain nucleus in the early stages of development.本研究采用鸟枪法蛋白质组学（shotgun-based proteomics）技术，对两个栽培小麦品种——普通小麦（*Triticum aestivum*，六倍体小麦，基因组组成为A、B、D）与一粒小麦（*T. monococcum*，二倍体小麦，基因组组成为A）的籽粒在两个早期发育阶段的核蛋白质组开展了分析。研究优化了细胞核纯化流程，并开发了改良的蛋白质样品制备方案，以高效去除淀粉与核酸等非蛋白杂质。最终共鉴定得到797种蛋白质，对应528种唯一蛋白质，其中36%被归入与DNA和RNA代谢相关的功能类别。此外还发现了107种功能未知蛋白及假定蛋白（hypothetical proteins）。部分鉴定获得的蛋白质可能具备多重功能，并可呈现多种亚细胞定位。基于串联质谱（MS/MS）分析结果，在两个物种及两个发育阶段中均检测到368种蛋白质，同时还发现了物种间与发育阶段间存在的部分定性差异。上述所有数据均阐明了籽粒细胞核在早期发育阶段的动态功能特性。