Kaspar-Schoenefeld Stephanie, Merx Kathleen, Jozefowicz Anna Maria, Hartmann Anja, Seiffert Udo, Weschke Winfriede, Matros Andrea, Mock Hans-Peter
Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Stadt Seeland OT Gatersleben, Germany.
Fraunhofer IFF, Magdeburg, Germany.
J Proteomics. 2016 Jun 30;143:106-121. doi: 10.1016/j.jprot.2016.04.007. Epub 2016 Apr 14.
Due to its importance as a cereal crop worldwide, high interest in the determination of factors influencing barley grain quality exists. This study focusses on the elucidation of protein networks affecting early grain developmental processes. NanoLC-based separation coupled to label-free MS detection was applied to gain insights into biochemical processes during five different grain developmental phases (pre-storage until storage phase, 3days to 16days after flowering). Multivariate statistics revealed two distinct developmental patterns during the analysed grain developmental phases: proteins showed either highest abundance in the middle phase of development - in the transition phase - or at later developmental stages - within the storage phase. Verification of developmental patterns observed by proteomic analysis was done by applying hypothesis-driven approaches, namely Western Blot analysis and enzyme assays. High general metabolic activity of the grain with regard to protein synthesis, cell cycle regulation, defence against oxidative stress, and energy production via photosynthesis was observed in the transition phase. Proteins upregulated in the storage phase are related towards storage protein accumulation, and interestingly to the defence of storage reserves against pathogens. A mixed regulatory pattern for most enzymes detected in our study points to regulatory mechanisms at the level of protein isoforms.
In-depth understanding of early grain developmental processes of cereal caryopses is of high importance as they influence final grain weight and quality. Our knowledge about these processes is still limited, especially on proteome level. To identify key mechanisms in early barley grain development, a label-free data-independent proteomics acquisition approach has been applied. Our data clearly show, that proteins either exhibit highest expression during cellularization and the switch to the storage phase (transition phase, 5-7 DAF), or during storage product accumulation (10-16 DAF). The results highlight versatile cellular metabolic activity in the transition phase and strong convergence towards storage product accumulation in the storage phase. Notably, both phases are characterized by particular protective mechanism, such as scavenging of oxidative stress and defence against pathogens, during the transition and the storage phase, respectively.
由于大麦作为全球重要的谷类作物，人们对确定影响大麦籽粒品质的因素高度关注。本研究聚焦于阐明影响籽粒早期发育过程的蛋白质网络。采用基于纳升液相色谱的分离技术与无标记质谱检测相结合的方法，以深入了解五个不同籽粒发育阶段（从预贮藏阶段到贮藏阶段，开花后3天至16天）的生化过程。多变量统计揭示了在分析的籽粒发育阶段存在两种不同的发育模式：蛋白质在发育中期（过渡阶段）或发育后期（贮藏阶段）表现出最高丰度。通过应用假设驱动的方法，即蛋白质免疫印迹分析和酶活性测定，对蛋白质组学分析观察到的发育模式进行了验证。在过渡阶段观察到籽粒在蛋白质合成、细胞周期调控、抗氧化应激防御以及通过光合作用产生能量方面具有较高的总体代谢活性。在贮藏阶段上调的蛋白质与贮藏蛋白积累有关，有趣的是，还与贮藏储备抵御病原体有关。在我们的研究中检测到的大多数酶的混合调控模式表明在蛋白质异构体水平存在调控机制。
深入了解谷类颖果的早期籽粒发育过程非常重要，因为它们会影响最终的籽粒重量和品质。我们对这些过程的了解仍然有限，尤其是在蛋白质组水平上。为了确定大麦籽粒早期发育的关键机制，采用了一种无标记的数据非依赖型蛋白质组学采集方法。我们的数据清楚地表明，蛋白质要么在细胞化和向贮藏阶段转变（过渡阶段，开花后5 - 7天）期间表现出最高表达，要么在贮藏产物积累期间（开花后10 - 16天）表现出最高表达。结果突出了过渡阶段多功能的细胞代谢活性以及贮藏阶段向贮藏产物积累的强烈趋同性。值得注意的是，这两个阶段分别以特定的保护机制为特征，即在过渡阶段清除氧化应激，在贮藏阶段抵御病原体。
