Analysis of regulatory networks underlying morphogenesis of leaf epidermis pavement cells

多细胞生物体从受精的卵细胞发展成具有特殊组织和器官的复杂生物体，需要精确的生长协调。驱动生长的基本过程是细胞分裂和细胞扩张，它们分别产生新的细胞和增加单个细胞的大小。在细胞扩张的过程中，细胞获得各种不同的形状来完成它们的特殊功能(细胞分化)。在植物中，细胞的扩张受到一个坚硬的细胞壁的控制和限制，该细胞壁将单个细胞包裹在质膜之外，并在物理上粘合相邻的细胞。细胞壁物质的沉积是由植物细胞骨架介导的，细胞骨架由微管和肌动蛋白细丝组成，形成一个高度动态的细胞内网络。细胞壁和细胞骨架的协同活动由复杂的信号网络协调，这些网络整合了内源和外部信号，以精确调控生长。叶表皮铺面细胞是植物中最复杂的细胞形态之一，在其背斜壁上有多个裂片和凹痕。除了对叶片生长的重要性和对光合作用活跃的组织的保护作用外，路面细胞因此是在机械耦合细胞组织的背景下研究多极生长的流行和合适的模型系统。遗传和药物研究，主要是在模式植物拟南芥中，已经确定了几个控制路面细胞形态发生的因素。然而，形状形成的确切机制和原则在很大程度上仍然是个谜。在我们之前的工作中，我们发现了一类植物特有的微管相关蛋白IQ67 DOMAIN5(IQD5)，作为一种新的路面细胞形状和细胞壁组成的调节因子，它可能整合了微管细胞骨架上的钙信号。此外，我们还开发了一个软件工具，首次实现了从显微镜图像中对路面细胞形状进行全自动检测和比较定量分析。我们首次对自然发生的拟南芥生态型中的路面细胞形状进行了分析，发现了很大的种内变异，这为识别相关的遗传位点提供了合适的基础。在拟议的项目中，我们的目标是利用细胞生物学和反向遗传学方法，从功能上表征IQD5在微管组织调控和细胞信号通路中的作用。此外，我们将使用正向遗传学方法，通过将拟南芥材料中的基因变异和表型变异联系起来，并通过筛选突变集合来识别新的调控因子。总之，通过反向遗传学和正向遗传学方法对路面细胞形状调节的联合分析将为研究细胞形态发生的机制提供一个合适的框架，并将有助于更全面地理解潜在的原理。

The development of multicellular organisms from a fertilized egg cell to complex organisms with specialized tissues and organs requires precise coordination of growth. The basic processes that drive growth are cell division and cell expansion, which generate new cells and increase the size of individual cells, respectively. During cell expansion, cells acquire various distinct shapes to fulfill their specialized functions (cell differentiation). In plants, cell expansion is controlled and restricted by a rigid cell wall that surrounds individual cells outside of the plasma membrane and physically glues neighboring cells. Deposition of cell wall material is mediated by the plant cytoskeleton, comprised of microtubules and actin filaments, which form a highly dynamic intracellular network. The concerted activity of the cell wall and the cytoskeleton is coordinated by complex signaling networks that integrate endogenous and external signals for precise regulation of growth. Leaf epidermis pavement cells form one of the most complex cell shapes in plants with multiple lobes and indentations in their anticlinal walls. In addition to their importance for leaf growth and their protective role for photosynthetically active tissues, pavement cells thus are a popular and suitable model system to study multipolar growth in the context of a tissue of mechanically coupled cells. Genetic and pharmaceutical studies, mostly in the model plant Arabidopsis thaliana, have identified several factors that control pavement cell morphogenesis. The precise mechanisms and principles underlying shape formation, however, remain largely enigmatic. In our previous work we identified a member of a plant-specific class of microtubule-associated proteins, IQ67 DOMAIN5 (IQD5), as novel regulator of pavement cell shape and cell wall composition, which likely integrates calcium signaling at the microtubule cytoskeleton. In addition, we have developed a software-tool, which for the first time enables fully automatic detection and comparative quantitative analysis of pavement cell shape from microscopy images. Our first analyses of pavement cell shape in naturally occurring Arabidopsis ecotypes revealed large intraspecific variation that provide a suitable basis to identify associated genetic loci. Within the proposed project we aim to functionally characterize the role of IQD5 in regulation of microtubule organization and in cellular signaling pathways, using cell biology and reverse-genetics approaches. In addition, we will use forward-genetics approaches to identify novel regulators by linking genotypic and phenotypic variation within Arabidopsis accessions and by screening of mutant collections. Collectively, the combined analysis of pavement cell shape regulation by reverse- and forward-genetics approaches will provide a suitable framework to investigate mechanisms of cell morphogenesis and will contribute to a more holistic understanding of the underlying principles.