Investigation of cell polarity mechanisms connected to cell fate and pattern formation in Arabidopsis vascular and stomatal development

植物是具有许多特殊组织的复杂有机体。细胞多样性的重要来源是不对称分裂的干细胞，这些干细胞产生不同命运的子细胞。为了定位和执行植物细胞谱系中的不对称分裂，细胞的极性是通过分子调控因子的极性定位来建立的，其作用模式仍然知之甚少。在这里，我将研究细胞极性蛋白是如何1)在不同的细胞谱系中调节细胞分裂和命运，2)在命运转换过程中以时空方式调节的，以及3)组织质膜蛋白质组/蛋白质组成的亚域。为了解决这些长期存在的问题，我将比较两种不同的、但在功能上相互依赖的、必不可少的植物结构的发育过程中所使用的极性机制：能够与环境进行气体交换的气孔和在整个植物中分配水分、营养物质和糖的维管系统。我的初步数据表明，打破气孔谱系中的不对称(BASL)是一种极性蛋白质，也是气孔谱系的基本调节因子，也调节维管细胞的分裂。利用遗传学和组织学的方法，我将进一步研究BASL的遗传框架及其与极地定位的血管调节因子的相互作用，如Brivis Radix(BRX)。另一个尚未探索的重要方面是对特定细胞类型内BASL蛋白水平的时空控制，我将使用一个强大的细胞培养系统来鉴定BASL降解机制的组件。我的第三种方法需要开创一种发育植物生物学的新技术--邻近标记--绘制和比较气孔和维管束中两个不同的极化亚域的质膜蛋白质组。使用特定的极性标记BASL-BRX和我确定的一种新的蛋白质来定位BRX结构域的对立面(抗BRX)，结合在宿主实验室成功建立的基于生物素的邻近标记方法，使我在确定极化质膜蛋白质组方面处于独特的地位。我希望对细胞的极性和命运产生新的见解，并特别揭示气孔和维管发育之间的细胞类型特定差异。由于气孔和维管系统从根本上决定植物的适合性，识别调节发育的共同极性机制不仅可以成为我独立研究生涯的起点，而且还可以帮助提高未来的作物产量。

Plants are complex organisms with many specialized tissues. Important origins of cellular diversity are asymmetrically dividing stem cells that produce daughter cells of different fate. To orient and execute asymmetric divisions in plant cell lineages, cell polarity is established by polar localization of molecular regulators whose modes of action are still only poorly understood. Here I will investigate how cell polarity proteins 1) regulate cell division and fate within distinct cell lineages, 2) are regulated in a spatio-temporal manner during fate transitions and 3) organize subdomains of the plasma membrane proteome/protein composition. To address these long-standing issues, I will compare polarity mechanisms used in the development of two distinct, but functionally co-dependent and essential plant structures: stomata that enable gas exchange with the environment and the vasculature that distributes water, nutrients and sugars throughout the plant. My preliminary data suggests that BREAKING ASYMMETRY IN THE STOMATAL LINEAGE (BASL), a polarity protein and fundamental regulator of the stomatal lineage, also regulates vascular cell divisions. Using genetic and histological approaches, I will further investigate the genetic framework of BASL and its interaction with polar localized vascular regulators, such as BREVIS RADIX (BRX). Another important unexplored aspect is the spatio-temporal control of BASL protein levels within specific cell types, and I will use a powerful cell culture system to identify components of BASL’s degradation machinery. My third approach requires pioneering a technique new to developmental plant biology - proximity labeling - to map and compare plasma membrane proteomes of two distinct, polarized subdomains in both stomata and vascular lineages. Using specific polarity markers BASL-BRX and a new protein I identified to localize opposite of the BRX domain (anti-BRX) in combination with a biotin-based proximity labeling approach successfully established in the host lab, puts me in a unique position to determine polarized plasma membrane proteomes. I expect to generate novel insights into cell polarity and fate in general, and to reveal cell-type specific differences between stomatal and vascular development in particular. As stomata and vasculature fundamentally determine plant fitness, identifying common polarity mechanisms regulating development could not only be the starting point of my independent research career but could also help to increase crop yield in the future.