花粉的萌发和随后的顶端生长是种子植物有性生殖的重要生理过程。这一过程与微丝骨架的动态变化密切相关。根据已有的研究推测，已发现和鉴定的植物微丝结合蛋白可能参与花粉管微丝骨架的动态调节。Villin/gelsolin/fragmin超家族蛋白是目前已知的唯一一类Ca2+依赖性肌动蛋白结合蛋白，其在植物细胞微丝骨架调控的过程中起着十分重要的作用。本研究对Villin/gelsolin/fragmin超家族蛋白LdABP41的功能进行了进一步深入的研究。首先，我们在本实验室先前工作的基础上对LdABP41与肌动蛋白的相互作用进行了进一步的研究。利用荧光显微镜观察发现，LdABP41具有Ca2+依赖的抑制单体肌动蛋白聚合过程的作用，使形成的actin主要以短而碎的小片段结构存在；LdABP41具有Ca2+依赖的剪切F-actin的作用, 使已聚合好的微丝片段化，微丝长度缩短。利用pyrene标记的actin进行微丝延伸实验和解聚实验，发现LdABP41具有Ca2+的结合微丝正端并抑制微丝正端延伸的功能；LdABP41也具有抑制微丝解聚的作用。这些体外结果表明LdABP41的存在使微丝结构维持在短小的微丝片段的状态。我们通过显微注射实验观察发现：LdABP41以浓度依赖的方式显著的缩短花粉管透明区长度并且减慢胞质环流的速度；通过二次注射的方法注入Alexa488-phalloindin标记花粉管中的微丝，发现其中有序的微丝结构受到破坏，最顶端的透明区开始出现短小的微丝片段，主干部位有序排列的微丝束遭到破坏，取而代之的是较为细长且不稳定的微丝，且排列的较为无序，另外主干部位也出现了较多短小的微丝片段。由体内实验结果结合体外实验的结果，我们推测LdABP41在体内可能通过片段化作用调节体内动态的微丝骨架并进一步影响花粉管的生长。我们通过免疫印迹实验观察了花粉发育各个阶段的蛋白表达水平。结果显示：在花粉的成熟阶段和花粉的花粉管生长阶段，蛋白表达水平基本保持一致；而在萌发过程中蛋白的表达水平出现了显著的变化：LdABP41在未水合和水合的花粉中大量存在，一旦萌发蛋白表达开始呈现显著的减少，并且随着萌发的过程这种减少不断加剧，与之相对应的微丝结构也发生了很大的转变。由这些结果，我们推测LdABP41在花粉的萌发过程中通过片段化的作用对微丝的动态结构起着重要的调控作用。

The sexual reproduction of spermatophyte depends on the germination and subsequent tip growth process of pollen, which correlates with the dynamic actin organization. Based on the previous research, it’s suggested that the actin binding proteins identified in the plant is likely involved in the dynamic regulation of actin structure. Villin/gelsolin/fragmin superfamily proteins are the only known Ca2+-dependent actin binding proteins, and have recently been shown to play important roles in tip growing plant cells. Present study intends to perform the further research on the identified member of villin/gelsolin/framin superfamily－LdABP41, and discuss the potential regulation mechanism of LdABP41 on the pollen germination process. According to the Ca2+-sensitive character of gelsolin superfamily, a novel Ca2+-dependent actin-binding protein with molecular mass of 41 kDa from lily pollen （Fan et al, 2004） was isolated and purified with DNase I chromatography. We performed further study on the in vitro and in vivo function of LdABP41 based on the previous work. Fluorescence microscope observation revealed that LdABP41 could inhibit G-actin polymerization in a Ca2+-dependent manner and lead to the formation of very short actin fragments in the field. LdABP41 also could sever populations of prepolymerized F-actin and shorten the F-actin length. In the actin seed elongation experiment, LdABP41 could cap the barbed end of the actin seed and prevent the F-actin elongation from barbed end in Ca2+-sensitive manner. Also LdABP41 could accelerate the F-actin depolymerization. It’s suggested that LdABP41 could maintain the actin array in the form of short F-actin fragment in vitro.Microinjection of LdABP41 into the growing pollen tube, it’s observed that LdABP41 could shorten the clear zone length and slow down the rate of cytopalsmic streaming in a concentration-dependent manner. Actin filament labeled with Alexa488-phalloidin revealed that the regular actin organization was disrupted; the short actin fragments appeared in the extreme tip region and also in the shank region. The parallel actin bundles in the shank region were instead by the thin and unstable actin filaments. In view of the in vitro and in vivo results, it’s deduced that LdABP41 is likely involved in the regulation of actin dynamics via fragmenting actin filaments in pollens. The dynamic LdABP41 expressing level was observed during the different phase of pollen development. The data showed that the expressing level of LdABP41 was maintained in constant level in the pollen maturing and the pollen tube growth period. However, the dynamic LdABP41 level suffered great change in the pollen germination process. Our results showed that a large amount of LdABP41 existed in the dehydrated and hydrated pollen. Once culturing in germination medium, the protein level decreased significantly, and the amount of LdABP41 was much less as the pollen germination process was going on. Correspondingly, the actin organization in the pollens suffered great change. According to the above results, we suggested that LdABP41 could play an important role in the regulation of actin dynamics during pollen germination via fragmenting actin filaments.