Structure-function analysis of the interaction of secretory peptides with their target proteins

在高等植物中，精子细胞是不能移动的。为了成功受精，它们需要通过花粉管的生长被输送到雌配子体。当到达珠孔时，花粉管停止生长并破裂，从而将精子细胞输送到卵器。在此背景下，我们最近发现类防御素(DEFL)蛋白ZmES4(玉米胚囊4)通过开放钾通道KZM1来介导玉米花粉管的破裂。由于这种相互作用的确切分子机制及其与花粉管破裂的关系尚不清楚，因此该提议的主要目的在于了解ZmES4与钾离子通道KZM1相互作用的结构-功能关系。由于这种相互作用是非常特定的，我们计划确定离子通道中对ZmES4结合至关重要的结构域或残基。此外，我们旨在阐明ZmES4介导的通道调节最终是如何导致花粉管破裂的。我们推测，除了K+外流，蔗糖吸收和随后的水内流是花粉管破裂的关键步骤。因此，我们将研究相应的蔗糖转运蛋白是否也受到ZmES4的影响。最后，如果时间允许，我们将阐明DEFL蛋白与离子通道的相互作用是否代表高等植物诱导花粉管破裂和精子释放的一般原理。因此，我们将测试DEFL蛋白和其他植物物种的离子通道，以了解它们履行这一功能的能力。在阳性检测的DEFL蛋白及其各自的靶标的情况下，我们将对它们的结构进行建模和比较，以澄清DEFL/靶标结合部位是否具有物种特异性结构。

In higher plants sperm cells are immobile. For a successful fertilization they need to be transported to the female gametophyte via growth of the pollen tube. Upon arrival at the micropyle the pollen tube stops to grow and bursts, thereby delivering the sperm cells to the egg apparatus. In this context, we have recently shown that the defensin-like (DEFL) protein ZmES4 (Zea mays embryo sac 4) mediates pollen tube burst in maize via opening of the potassium channel KZM1. Since the exact molecular mechanism of this interaction and its relation to pollen tube burst is not known a main goal of the proposal relies on understanding the structure-function relationship of the interaction of ZmES4 with the potassium ion channel KZM1. As this interaction is very specific, we plan to identify domains or residues within the ion channel which are essential for ZmES4 binding. Furthermore we aim on elucidating how ZmES4-mediated channel modulation finally leads to pollen tube burst. We postulate that in addition to K+ flux, sucrose uptake and ensuing water influx represents an essential step for pollen tube rupture. Therefore we will investigate whether the corresponding sucrose transporters are also affected by ZmES4 application. Finally, if time permits we will elucidate whether the interaction of DEFL proteins with ion channels represents a general principle in higher plants to induce pollen tube rupture and sperm release. Therefore, we will test DEFL proteins and ion channels of other plant species regarding their ability to fulfill this function. In case of positively tested DEFL proteins and their respective targets we will model and compare their structures to clarify if the DEFL/target binding site possesses a species-specific structure.