RhoGTPase signalling in double fertilisation in Arabidopsis and maize (A15)

双受精是所有开花植物种子生产的关键过程，因此对作物生产至关重要。在双受精过程中，两个雄性精子细胞与两个雌性配子融合，分别发育成胚胎和滋养胚乳。该过程的一个基本特征是花粉管的尖端定向生长穿过花组织并将雄配子递送给雌配子体（Bleckmann et al.，2014; Dresselhaus等人，2016）。这种极尖生长需要多个反馈回路来平衡细胞扩张和稳定性。植物的RhoGTP酶RHO（ROP）是整合这些反馈回路的核心因素。它们的活性通过激活ROP鸟嘌呤核苷酸交换因子（ROPGEF）和失活GTP酶激活蛋白（GAP）来调节（Qin和Yang，2011）。在以前的研究中，只研究了个别蛋白质或机制。然而，我们和其他人发现，这些因子中的多个虽然在同一细胞中表达，但它们是一致或按顺序起作用的。此外，我们发现不同的ROPGEF调节相同ROP在根毛中的定位或活性，根毛是植物中的第二极尖生长过程（Denninger等人，2019年）。基于这些观察不同的ROPGEF功能在根毛，我假设ROP信号的不同方面受到不同的ROPGEF在花粉管。更好地了解ROPGEFs在花粉管生长过程中的作用将有助于我们更好地理解种子生产。在本项目中，我们将研究三种优先在花粉中表达的ROPGEFs在拟南芥（Arabidopsis thaliana）花粉管顶端生长和配子相互作用中的具体功能。我们的目标是确定他们的特定功能ROP定位和激活检查钙离子信号和肌动蛋白组织读出。为了研究植物之间的这些机制的保护，我们将调查ROPGEF功能在玉米花粉管中使用敲除和敲低突变体，并比较这ROPGEF功能在拟南芥。此外，我们将研究ROP信号在雌性拟南芥配子体，这条途径仍然完全未被探索。初步结果支持至少一个ROPGEF在该组织中的表达，我们将进一步研究其他ROP信号传导因子的存在，以及它们在双受精过程中的雌配子体细胞中的功能。

Double fertilisation is a key process for seed production in all flowering plants and therefore crucial for crop production. During double fertilisation, two male sperm cells fuse with two female gametes, which develop into the embryo and the nurturing endosperm, respectively. An essential feature of this process is that the tip of the pollen tube grows directionally through the floral tissue and delivers male gametes to the female gametophyte (Bleckmann et al., 2014; Dresselhaus et al., 2016). This polar tip growth requires multiple feedback loops for balanced cellular expansion and stability. The RhoGTPases RHO OF PLANTS (ROPs) are central factors to integrate these feedback loops. Their activity is regulated by activating ROP guanine nucleotide exchange factors (ROPGEFs) and inactivating GTPase-Activating Proteins (GAPs) (Qin and Yang, 2011). In previous studies, only individual proteins or mechanisms were investigated. We and others, however, showed that multiple of these factors, although expressed in the same cell, act in concert or in sequence. Additionally, we found that distinct ROPGEFs regulate either localisation or activity of the same ROP in root hairs, a second polar tip growth process in plants (Denninger et al., 2019). Based on these observations of distinct ROPGEF functions in root hairs, I hypothesise that different aspects of ROP signalling are regulated by different ROPGEFs in pollen tubes. A better understanding of ROPGEFs during pollen tube growth would enhance our understanding of seed production.In this project, we will investigate the specific functions of three, preferentially pollen-expressed ROPGEFs in pollen tubes during tip growth and gamete interaction in Arabidopsis thaliana. We aim to identify their specific functions on ROP localisation and activation by examining Ca2+ signalling and actin organisation as readouts. To study the conservation of these mechanisms between plants, we will investigate ROPGEF function in pollen tubes of Zea mays using knock-out and knock-down mutants and compare this to ROPGEF function in Arabidopsis. Additionally, we will investigate ROP signalling in the female Arabidopsis gametophyte, where this pathway remains completely unexplored. Preliminary results support expression of at least one ROPGEF in this tissue, and we will further investigate the presence of other ROP signalling factors, as well as their function in cells of the female gametophyte during double fertilisation.