腹毛类原生动物的射出胞器研究

Extrusive organelles are typical dense core granules that locate in cell cortex of various protozoans. Studying these special organelles is of great significance in further understanding the organelle evolution in protozoa and cell regulation in eukaryotic cells. The proposed project will carry out researches on various taxa and different physiological states of the highly differentiated hypotrichous protozoans to reveal the ultrastructure, composition and function of extrusomes: using scanning and transmission electron microscopy as well as enzyme cytochemistry methods to reveal the basic ultrastructure and diversity of these extrusive organelles and define the morphologically so-called "cortical granules”, meanwhile, obtain the development and emission mechanisms as well as the differentiation characteristics in special cell physiological states; using two-dimensional electrophoresis, chromatography, mass spectrometry and other related technologies to detect the components, and functional or specific proteins in the extruded structure of typical extrusomes; using experimental ecological methods to test the functions and action mechanisms of typical extrusomes. The result generated in the project will provide new information for understanding the complexity and functions of extrusomes in protozoans, as well as the mechanisms of cellular structure formation and protein level cell regulation.

射出胞器是一类广泛存在于各类原生动物细胞皮层下的致密核心型囊泡。研究此类胞器对于进一步理解原生动物各阶元细胞器水平的演化规律及真核细胞的细胞调控机制具有重要理论价值。本项目拟以纤毛器高度分化的腹毛类原生动物不同种类、各种生理状态下的细胞为主要研究材料，从此类皮层细胞器的基本超微结构、物质构成、胞器功能三个不同角度开展系统研究：应用扫描、透射电镜术及酶细胞化学术等手段揭示射出胞器的基本超微结构特征及类群中的多样性、给出所谓“皮层颗粒”的细胞学定义，同时探索射出胞器在细胞内发生、发射机制以及在特殊生理时期细胞中的分化特征；应用双向凝胶电泳及色谱、质谱等技术检测典型射出胞器射出结构的物质组分和功能性、特异性蛋白类型；利用实验生态手段获取典型腹毛类射出胞器的功能及作用机制。所获结果可为揭示原生动物射出胞器的复杂性和功能、原生动物细胞结构形成和蛋白水平调控的机理提供新的资料。

射出胞器是一类广泛存在于各类原生动物细胞皮层下的致密核心型囊泡。本项目针对广义腹毛类这一高等类群的射出胞器，利用显微及亚显微形态观察、胞器分离纯化和质谱检测等手段从不同角度开展研究。主要结果包括：（1）刻画了19种腹毛类的射出胞器的形态，并主要依据超微结构将它们归纳为7个类型；同时发现射出胞器类型与物种基于SSU rRNA基因的系统发育关系相吻合，因而可作为腹毛类分类鉴定和系统修订的新依据。（2）揭示了色素泡、类黏液泡和类色素泡均起源于含有内质网及线粒体的细胞质囊泡，且均经历了囊泡内发育过程；但不同类型射出胞器在休眠期的命运不同。（3）从形态学角度揭示了类刺丝泡在发射机制上不同于其它已知类型，即发射时经历帽部解体、体部二次展开和中央轴杆推出过程。（4）通过检测蛋白成分，发现类刺丝泡和角状黏液泡的发射可能经历钙离子介导的膜融合并且是一个耗能的过程；这两种射出胞器的形成过程涉及细胞内囊泡合成与转运和内质网-高尔基体蛋白转运相关通路。本研究为原生动物疑难阶元的系统分类提供了新视角，也为探讨细胞结构与功能及真核细胞的演化等问题提供了新资料。此外，依托该项目，已发表文章16篇（含12篇SCI文章），培养5名研究生。