Rotation 1: Validation of a putative MYB transcription factor involved in chloroplast development

第 1 轮：验证参与叶绿体发育的推定 MYB 转录因子

• 批准号: 2887717
• 金额: --
• 依托单位: University of Cambridge
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2023
• 资助国家: 英国
• 起止时间: 2023 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2887717
• 关键词: Rotation; Validation; putative; MYB; transcription

BBSRC strategic theme: Bioscience for sustainable agriculture and foodWheat is crucial to UK agriculture (https://www.gov.uk/government/statistics/agricultural-land-use-in-the-united-kingdom/agricultural-land-use-in-united-kingdom-at-1-june-2023). The circadian oscillator regulates several pathways underlying yield-related traits, including heading date and temperature response (Asseng et al., 2015; Wittern et al., 2023). However, differences between wheat and the model plant Arabidopsis thaliana obstruct the application of our understanding of these pathways to crops. While CONSTANS regulates flowering time in A. thaliana, wheat heading date is determined by PHOTOPERIOD-1 (Ppd-1) and EARLY FLOWERING 3 (ELF3) (Alvarez et al., 2023; Shaw et al., 2020; Suárez-López et al., 2001). The mechanism of circadian oscillator regulation by temperature is also unclear in wheat; in A. thaliana, ELF3 has been proposed to respond to temperature through a predicted prion domain (PrD) that is not present in wheat ELF3 (Jung et al., 2020; Ronald et al., 2021; Zhu et al., 2023). A better understanding of the wheat circadian clock is thus crucial to breeding strategies targeting clock genes to improve the resilience of wheat to climate change (Steed et al., 2021).In this project, we propose to use molecular and biochemical methods to better understand the structure and function of the wheat circadian clock. To facilitate wheat chronobiology research, we plan to develop a bioluminescent clock gene reporter line to measure circadian rhythms at the genetic level. A reporter can then be crossed into clock gene mutant lines. In addition to this broader aim, we will focus on determining the role of ELF3 within the circadian oscillator. Firstly, we will assess whether an Evening Complex (EC) with LUX ARRHYTHMO (LUX) and EARLY FLOWERING 4 (ELF4) orthologs forms in wheat using in silico and in vivo assays (Herrero et al., 2012; Nusinow et al., 2011). Secondly, we will test the interaction of ELF3 with orthologs of partners from A. thaliana, such as TIMING OF CAB 1, CONSTITUTIVE PHOTOMORPHOGENIC 1, and GIGANTEA (Huang and Nusinow, 2016). To complement this work, we plan to build on ongoing work analysing the wheat circadian transcriptome by investigating the binding of ELF3 to target gene promoters using chromatin immunoprecipitation sequencing (ChIP-seq). We will also investigate the molecular mechanisms of yield-related circadian oscillator output pathways, including flowering time and thermomorphogenesis. In wheat, the regulation of flowering time involves Ppd-1, VERNALIZATION 1 (VRN1), VRN2, and VRN3 and responds to photoperiod and vernalization (Distelfeld et al., 2009). ELF3 integrates this pathway with the clock; we aim to determine whether this regulation occurs through an EC (Alvarez et al., 2023; Wittern et al., 2023). Additionally, thermo-responsive growth can be mediated independent of flowering time (Wang et al., 2024). We will therefore test the involvement of ELF3 through in vivo assays, potentially expanding this to test the broader ELF3 interactome using methods such as affinity purification-mass spectrometry (AP-MS) (Box et al., 2015; Huang and Nusinow, 2016). This work thus aims to elucidate the molecular mechanisms underlying the wheat circadian oscillator and its yield-related output pathways. This can enable the application of this research to agriculture, for example, in the form of breeding targets.

(https://www.gov.uk/government/statistics/agricultural-land-use-in-the-united-kingdom/agricultural-land-use-in-united-kingdom-at-1-june-2023).战略主题：生物科学促进可持续农业和食品小麦对英国农业至关重要昼夜节律振荡器调节产量相关性状的几个途径，包括抽穗期和温度反应(Asseng等人，2015；Wittern等人，2023)。然而，小麦和模式植物拟南芥之间的差异阻碍了我们对这些途径在作物上的应用。小麦抽穗期由光周期1(PPD-1)和早花3(ELF3)决定(Alvarez等人，2023年；Shaw等人，2020年；Suárez-López等人，2001年)。在小麦中，温度调节昼夜节律振荡器的机制也不清楚；在A.thaliana中，ELF3已被提议通过小麦ELF3中不存在的预测的Prion结构域(PRD)来响应温度(Jung等人，2020；Ronald等人，2021；朱等人，2023)。因此，更好地了解小麦生物钟对于针对时钟基因的育种策略至关重要，以提高小麦对气候变化的适应能力(Steed等人，2021)。在这个项目中，我们建议使用分子和生化方法来更好地了解小麦生物钟的结构和功能。为了方便小麦时间生物学的研究，我们计划开发一个生物发光钟基因报告系来在遗传水平上测量昼夜节律。然后，记者可以被杂交到时钟基因突变系中。除了这个更广泛的目标，我们将专注于确定ELF3在昼夜节律振荡器中的作用。首先，我们将评估小麦中是否存在与LUX ARRHYTHMO(LUX)和早花4(ELF4)同源的晚间复合体(EC)(Herrero等人，2012；Nusinow等人，2011)。其次，我们将测试ELF3与拟南芥伙伴的同源基因的相互作用，例如CAB 1、构成光形态发生1和巨型茶树的时机(Huang和Nusinow，2016)。为了补充这项工作，我们计划在正在进行的工作的基础上，通过使用染色质免疫沉淀测序(CHIP-SEQ)研究ELF3与靶基因启动子的结合来分析小麦昼夜节律转录组。我们还将研究与产量相关的昼夜节律振荡器输出途径的分子机制，包括开花时间和热形态发生。在小麦中，开花时间的调节涉及PPD-1、VRN1、VRN2和VRN3，并对光周期和春化做出反应(Distelfeld等人，2009)。ELF3将这一途径与时钟整合；我们的目标是确定这种调控是否通过EC发生(Alvarez等人，2023；Wittern等人，2023)。此外，温度反应生长可以独立于开花时间进行调节(Wang等人，2024)。因此，我们将通过体内分析来测试ELF3的参与，可能会扩大到使用亲和纯化-质谱分析(AP-MS)等方法来测试更广泛的ELF3交互作用组(Box等人，2015；Huang和Nusinow，2016)。因此，这项工作旨在阐明小麦昼夜节律振荡器及其产量相关输出途径的分子机制。这可以使这项研究能够应用于农业，例如，以育种目标的形式。