Cell-specific analysis of plant shade avoidance responses

描述(申请人提供)：植物为固着植物，其发育受到其环境的精确控制。作为对即将到来的阴影的反应，植物的结构经历了深刻的形态变化，包括茎的伸长和加速开花。这种表型在农业上有影响，在密植植物中的遮荫回避反应可能会减少可收获器官的生长。因此，干扰遮荫所需的信号通路可能会产生更好的产量。以前对模式植物拟南芥的研究表明，遮荫是信号信号，因为叶片中植物激素生长素的合成迅速增加。生长素随后被运输到茎中，以促进伸长生长。因此，很可能特定类型的细胞对遮荫有特殊的反应。然而，由于分离特定细胞群体所需技术的局限性，目前还缺乏对拟南芥新梢中细胞特异性表达变化的分析。在这里，最近发展的完整方法学将被用来研究不同类型的茎细胞中基因表达的变化。叶片和子叶感知许多光信号，这些器官的三个主要组织--叶肉、表皮和维管系统的核转录产物将被提纯。利用RNA测序技术，这项提议将测试这些细胞中基因表达的变化，以响应模拟阴影。通过将这些变化与遮荫诱导生长素生物合成缺陷的突变植物的变化进行比较，可以确定特定细胞类型中基因表达的变化是否部分位于生长素信号的下游。利用从这一分析中收集的信息，潜在有趣的细胞特定遮荫调节基因的可用T-DNA插入突变体将被测试遮荫回避表型的缺陷。有趣的候选基因可能是生长素生物合成基因，它可能是空间控制的，或者转录调节基因，其中一些已经被认为在遮荫避免表型中起作用。将通过在相关的细胞特异性启动子下表达该基因，并测试这如何影响植物的环境反应，来评估该基因在调节植物结构中的细胞特异性作用。这一分析将把遮荫避免所需的新颖和先前确定的因素置于空间、细胞类型的特定背景中。最终，这里确定的基因的空间扰动可能会产生植物，在这些植物中，对阴影的反应结构变化是以器官特有的方式控制的。这在商业植物物种中可能是有利的，在商业植物物种中，遮荫反应可以在空间上进行调整，以在给定的环境中产生最大产量。此外，该提案试图了解环境变化是如何在细胞水平上传递的，以影响有机体在发育过程中的身体计划。

DESCRIPTION (provided by applicant): Being sessile, a plant's development is exquisitely controlled by its environment. In response to impending shade, a plant's architecture undergoes a profound morphological change, including stem elongation and accelerated flowering. This phenotype has consequences in agriculture, where shade avoidance responses in densely grown plants may reduce the growth of harvestable organs. As such, the perturbation of signaling pathways required for shade avoidance may produce crops with better yields. Previous studies of the model plant Arabidopsis thaliana have shown that shade signals because a rapid increase in the synthesis of the phytohormone auxin in leaves. Auxin is subsequently transported to stems to promote elongation growth. Therefore, it is likely that specific cell types have specialized responses to shade. However, the analysis of cell-specific expression changes in the Arabidopsis shoot has been lacking, primarily due to limitations in techniques required to isolate a given cell population. Here, the recently developed INTACT methodology will be used to study gene expression changes in shoot cell types. The leaf and cotyledons sense many light signals, and nuclear transcripts from the three major tissues of these organs, the mesophyll, epidermis and vasculature, will be purified. Using RNA-Sequencing technology, this proposal will test the changes in gene expression in these cells in response to simulated shade. By comparing these changes with that of mutant plants defective for shade-induced auxin biosynthesis, it will be determined if the gene expression changes in a given cell type are in part downstream of auxin signals. Using the information gleaned from this analysis, available T-DNA insertion mutants of potentially interesting cell-specific shade-regulated genes will be tested for defects in shade avoidance phenotypes. Interesting candidate genes would be auxin biosynthesis genes, which may be spatially controlled, or transcriptional regulators, some of which are already known to have a role in the shade avoidance phenotype. The gene's cell-specific role in modulating plant architecture will be assessed by expressing the gene under relevant cell-specific promoters and testing how this affects plant environmental responses. This analysis will place novel and previously identified factors required for shade avoidance into a spatial, cell-type specific context. Ultimately, the spatial perturbation of the genes identified here may generate plants where architectural changes in response to shade are controlled in an organ-specific manner. This may be advantageous in commercial plant species, where shade responses can be spatially tuned to produce maximum yield in a given environment. Furthermore, the proposal seeks to understand how environmental change is transduced at the cellular level to affect an organism's body plan during development.