Transcriptional landscaping in plant stem cells: from chromatin to gene regulatory networks

所有陆地生命最终都依赖于植物分生组织--一小群未分化的细胞，它们产生所有主要的植物器官，如叶子和花。在拟南芥等高等植物的顶端分生组织(SAM)中，基因调控网络(GRN)控制细胞的生长、命运和特性，以平衡新器官的形成和分生组织的完整性，确保生长所需的多能细胞的可持续供应。转录因子，特别是同源结构域蛋白芽分化蛋白(STM)，通过调控靶基因的表达，在SAM的功能中发挥关键作用。然而，对STM GRN的组成和结构还没有完全了解。本项目旨在对SAM中的STM基因调控网络有一个全面的了解。STM编码一种仅在分生组织中表达的结节样同源结构域转录因子。STM功能的丧失导致不能发育或维持分生组织，而STM的过度表达抑制了叶片细胞的分化，促进了异位茎分生组织的从头形成。这些显着的表型变化表明，STM在调控分生组织发育和功能的GRN(S)中发挥着核心作用。为了了解STM是如何运作的，以及它的作用是如何与更广泛的分生组织GRN整合在一起的，将采用多层转录景观方法。首先，将使用可诱导的STM过表达和RNAi(基因沉默)以及全基因组RNA-SEQ分析来识别可能的STM靶基因。为了确定哪些STM反应基因直接受STM调控，将进行全球染色质免疫沉淀(CHIP-SEQ)以确定STM结合的基因组区域。由于STM还影响染色质结构以控制细胞的分化状态，染色质粒子光谱分析(CPSA)将用于分析STM结合的基因组区域的染色质结构，将假定的STM结合部位置于更广泛的核小体和其他DNA结合因子的范围内。然后将使用贝叶斯网络分析来推断GRN结构和STM目标基因之间的关系。该项目是一个令人兴奋的机会，可以在两位成熟的分生组织生物学专家的监督下进行培训，并学习多学科的技能和技术，包括分子遗传学、植物细胞和组织培养、生物信息学和发育生物学。该项目将为控制分生组织发育的网络带来新的见解，并可能导致操纵植物生长、结构和产量的新策略。

All terrestrial life ultimately depends on plant meristems - small groups of undifferentiated cells that produce all major plant organs such as leaves and flowers. In the shoot apical meristem (SAM) of higher plants such as Arabidopsis, gene regulatory networks (GRNs) control cell growth, fate and identity to balance the formation of new organs, such as leaves and flowers, with maintenance of meristem integrity, ensuring the sustainable supply of pluripotent cells necessary for growth. Transcription factors, particularly the homeodomain protein SHOOT MERISTEMLESS (STM), play critical roles in SAM function through regulation of target gene expression. However, the components and structure of the STM GRN are not fully understood. This project aims to develop a comprehensive understanding of the STM gene regulatory network in the SAM. STM encodes a Knotted1-like TALE homeodomain transcription factor that is expressed only in the meristem. Loss of STM function leads to failure to develop or maintain the meristem, while STM overexpression inhibits leaf cell differentiation and promotes the de novo formation of ectopic shoot meristems. These dramatic phenotypic changes suggest a central role for STM in the GRN(s) that regulate meristem development and function. To understand how STM operates and how its role is integrated with the wider meristem GRNs, a multi-layered transcriptional landscaping approach will be undertaken. First, putative STM target genes will be identified using inducible STM overexpression and RNAi (gene silencing) followed by genome-wide RNA-seq analysis. In order to determine which of these STM-responsive genes are directly regulated by STM, global chromatin immunoprecipitation (ChIP-seq) will be performed to identify genomic regions to which STM is bound. Since STM also affects the chromatin structure to control the differentiation status of cells, chromatin particle spectrum analysis (CPSA) will be used to analyse the chromatin structure in the genomic regions bound by STM, placing the putative STM binding sites within the wider landscape of nucleosomes and other DNA-bound factors. Bayesian network analysis will then be used to infer GRN structure and relationships among the STM target genes. This project is exciting opportunity to train under the supervision of two established experts in meristem biology, and to learn multidisciplinary skills and techniques including molecular genetics, plant cell and tissue culture, bioinformatics and developmental biology. The project will lead to new insights into networks controlling meristem development and could lead to novel strategies for the manipulation of plant growth, architecture and yield.