阿拉伯半乳糖蛋白(AGPs)调控油菜小孢子胚胎顶-基轴向分化的机制研究

Pattern formation of embryogenesis is a crucial event during the life cycle of higher plants,which originated from the fertilized egg cell resulted in the matured embryo through a series of cell division and differentiation.Arabinogalactan proeins represent a class of proteoglycans that is widespread throughout the plant kingdom.These molecules are invovled in the regulation of several important events related to plant growth and development.Previous work indicated that these proteins have important roles for initiating cell developmental fate and for maintaining embryonic normal development during the course of microspore embryogenesis in Brassica napus cv L. In this project,the system of isolated microspore embryogenesis, the combination of zygotic embryo in vitro culture system, is still used for research object. It will reveal that the regulatory mechanism of arabinogalactan proteins on cell division,cell differentiation and cell fate during the early pattern formation of embryogenesis,based on effect of these proteins on early embryonic apical-basal pattern formation of microspore-derived embryo through further analysis of characterization of the structure,temporal and spatial expression of arabinogalactan proteins recognited by different antibodies used a variety of research methods and stools. The reults will help us to have a new understanding how arabinogalactan proteins involve in pattern formation of embryo, and to provide a new evidence which will decrypt the issues related to embryogenesis.So It is very obvious for theoretical significance of this project.

高等植物胚胎发生模式建成一直是植物发育生物学的热点问题。它起始于受精的卵细胞，经过一系列精巧而复杂的细胞分裂与分化后发育成为成熟的胚。阿拉伯半乳糖蛋白（AGPs）是植物细胞质外体中的主要成分之一，它对植物的生长和发育起着重要的调控作用，目前越来越受到研究者的关注。前期工作表明，该蛋白在油菜小孢子胚胎发生过程中对胚的起始发育命运和维持胚的正常发育模式有重要的作用。本项目仍然以油菜小孢子离体胚胎发生系统为研究对象，结合合子胚离体培养系统，从不同抗体识别的AGPs在油菜小孢子胚胎发生过程中的时空表达特点和结构特征入手，结合多种研究方法和手段，深入分析阿拉伯半乳糖蛋白对小孢子胚胎发生早期顶-基模式建成的影响，揭示该蛋白在胚胎早期模式建成中对细胞分裂、细胞分化和细胞发育命运的调控机制。本项目的研究结果有助于我们对阿拉伯半乳糖蛋白参与胚胎模式建成有新的证据，其理论意义十分明显。

高等植物胚胎发生模式建成一直是植物发育生物学的热点问题，它起始于受精的卵细胞，经过一系列精巧而复杂的细胞分裂与分化后发育成成熟胚。阿拉伯半乳糖蛋白是植物细胞质外体中的主要成分之一，它对植物的生长和发育起重要的调控作用，目前越来越受到研究者的关注。本项目以油菜小孢子离体胚胎发生系统为研究对象，结合合子胚离体培养体系，从不同抗体识别的AGPs在油菜小孢子胚胎发生过程中的时空表达入手，结合多种研究方法和手段，深入分析了阿拉伯半乳糖蛋白对小孢子胚胎发生早期顶基模式建成的影响揭示了该蛋白在胚胎早期模式建成中对细胞分裂、细胞分化和细胞发育命运的调控机制。主要结果如下：.1.比较了两种离体胚胎发生体系中不同抗体识别的AGPs在胚胎发育中的时空表达，AGPs主要定位在小孢子胚的胚体上。用与AGPs专一结合的化学试剂 βGlc Y处理，导致胚胎发育顶基模式受到严重影响。.2.采用分子和生化技术，证实了βGlc Y处理影响了AGPs小孢子胚中蛋白的表达，通过RT-PCR进一步确证了βGlc Y影响胚胎发育相关的基因，进而导致顶基模式的改变；.3.利用转基因油菜，揭示了βGlc Y处理小孢子胚后改变了生长素的极性运输。.4.利用转基因油菜，揭示了βGlc Y处理后，小孢子胚表皮细胞发育命运的改变是由于相关胚性基因GRP、LEC1的表达下降引起。.5.初步探讨了胚体与胚柄之间的相互作用，验证了胚体对胚柄的胚性发育的抑制作用。.6.通过免疫组化技术，揭示AGPs可能作为信号分子影响小孢子在花药中的发育。