【作者】 李俊纲；

【导师】 顾福康；

【作者基本信息】 华东师范大学 ， 动物学， 2004， 硕士

【摘要】 为揭示纤毛虫在休眠条件下细胞结构于不同水平的分化及细胞生命活动特征，本文应用生化抽提及SDS-PAGE凝胶电泳、包埋—去包埋透射电镜技术、常规透射电镜技术等方法，对大尾柱虫(Urostyla grandis)的营养细胞和休眠细胞的超微结构、类中间纤维—核纤层—核骨架体系以及该体系的蛋白组分等进行了比较研究。 1．超微结构 大尾柱虫在形成休眠细胞后，所特化出的包囊壁在形态学上可分为三层，从外向内依次是外层壁、内层壁和颗粒层，纤毛杆和微管结构都已被皮层吸收，仅残留部分毛基体。在此基础上，本实验证实，在休眠细胞解脱过程中，包囊壁经历了一个逐渐变薄并最终断裂的过程，纤毛杆从残留的毛基体位置逐渐长出，口围带在休眠细胞解脱的后期形成。本实验还在大尾柱虫休眠细胞解脱过程中发现其自噬泡含有比较完整的拟糖原颗粒、线粒体和膜性结构，有的自噬泡周围有很多核糖体。在休眠细胞解脱过程中，内膜系统也变得逐渐发达，可能发育成内质网和高尔基体的网状结构也相应形成。在休眠细胞中大部分线粒体变得胀大、内外膜模糊、内部嵴也模糊，有时还会聚集，在解脱过程中，这些现象逐渐消失。通过对休眠细胞解脱过程中大核形态的大量观察，发现有的大核呈“哑铃形”，有的大核中间膨大，两端与中间部分有缢痕，这说明大核至少有两种分裂方式——二分裂和三分裂，大核很可能就是通过这两种分裂方式恢复其数量。小核在休眠细胞解脱过程中没有明显的形态学上的变化，它可能与休眠细胞的解脱过程没有联系。这些结果为进一步研究纤毛虫细胞休眠和解脱机制提供了资料。 2．类中间纤维—核纤层—核骨架体系及其蛋白组分 在大尾柱虫的营养细胞和休眠细胞内均存留有类中间纤维—核纤层—核骨架体系，该体系的整体结构和基本形态单元非常相似，构成此体系的基本形态单元是直径约为10nm的单根纤维或由单根纤维合并而成的纤维束。在细胞质中原来相当于细胞核的区域，由核纤层维持了原来的核区，核纤层的纤维联络核骨架纤维及位于细胞质中的类中间纤维，类中间纤维又终止于位于细胞周缘的、由多根纤维合并而成的支架上，三者之间形成有机的联系，在细胞内构成了一个连续的、稳定的三维网架体系。从形态学方面来讲，该体系在细胞中起支架作用并可能与各细胞器的定位有关，是保持细胞内结构完整性的主要因素。在纤毛虫中，类中间纤维—核纤层—核骨架体系的稳定性，暗示在纤毛虫细胞生命活动中，该体系可能起着相当重要的作用。2004届硕卜学位论文大尾柱虫背养细胞与休眠细胞超微结构和类中间纤维的比较研究 大尾柱虫在形成休眠细胞后，类中间纤维一核纤层一核骨架体系蛋白组分发生了一定的变化:营养细胞中上述骨架蛋白的电泳条带有29条，休眠细胞中该骨架蛋白的电泳条带有24条，其中营养细胞中特有的蛋白条带有4条，休眠细胞特有的蛋白条带有1条，休眠细胞与营养细胞共享的蛋白条带有23条。从蛋白组分来讲，休眠细胞中保留了大部分营养细胞中的类中间纤维一核纤层一核骨架体系蛋白组分，与电镜观察的该体系的整体结构和基本形态单元在营养细胞和休眠细胞中具有相似性这一结果一致;同时失去了营养细胞中的一些组分也产生了自身的一些独特组分，说明在纤毛虫休眠细胞中，类中间纤维一核纤层一核骨架体系蛋白组分的变化有可能反映了细胞休眠以后基因表达状态的改变。这些结果为深入研究细胞的代谢特征和调控机制奠定了基础，对揭示细胞生命活动的特征有重要意义。更多还原

【Abstract】 In order to reveal the different differentiation of cellular structures and characteristics on the condition of resting state in ciliates, the vegetative cell and resting cyst in Urostyla grandis were used as experimental materials. By using the methods of selective extraction, SDS-PAGE, DGD embedding and embedment-free transmission electron microscope (TEM) and TEM, a comparative study on ultrastructure and intermediate-type filament -lamina-nuclear matrix system and its protein compositions of the vegetative cell and resting cyst in Urostyla grandis were carried.1. UltrastructureAfter the formation of resting cyst in Urostyla grandis, the cyst wall is composed of three layers named from outside to inside: ectocyst, endocyst, and granular layer. Ciliary shaft and microtubule are absorbed by cortex , except for part of the kinetosome. It is proved that cyst wall undergo the process of becoming thinner and thinner and break at last, the ciliatures grow gradually from the position of kinetosomes and adoral zone of membranelle(AZM) is formed during the resting cyst’s differentiation . It is found that autophagic vacuoles include almost integrated paraglycodens, mitochondriae and membrane structures, some ones are attached around by many ribosomes, during the resting cyst’s differentiation. The inner membrane system becomes complex gradually, The net structure possibly developing into endoplasmic reticulum and Golgi body, form accordingly, during the resting cyst’s differentiation. A lot of mitochondriae become bigger, its outer and inner membrane become vague, the cristae in them also become vague, sometimes they will converge, in the resting cyst, these phenomena will disappear during the resting cyst’s differentiation. With a great of observation to the macronuclear’s shapes, It is obtained that some macronuclears display the shape of dumb-bell, some macronuclears expand in their middle and become thinner between middle and two ends. So the macronuclears perhaps have two ways of divisions - two divisions and three divisions at least, It is possible that the macronuclears recover their number using the two ways. No obvious changes happen in the micronuclear’s shape, which perhaps have no relation with the process of resting cyst’s differentiation. These results provide documents for more study on the mechanism of cellular resting and differentiation in ciliates.2. Intermediate-type filament-lamina-nuclear matrix system and its protein compositionsThe intermediate-type filament-lamina-nuclear matrix system is left in the vegetative cell and resting cyst of Urostyla grandis, whose entirety structure andbasic shape unit are similar, and whose basic shape unit is made up with single filaments that diameter is 10 nm or with filament bundle that are combined by single filaments. The area equal to the nuclear in the cell, is maintained by the lamina, while the filaments of the lamina connect the nuclear matrix filaments and the intermediate-type filaments in cytoplasm(CIFs), and the CIFs end at the scaffold which locate at the edge of cell and are combined with many filaments. The three kinds of filaments make an organic entirety, and build up a continue and stable net system in cell. Talking it from the morphology, the net system plays a role of frame, and is concerned with the cellular organs location, which is also an important factor to keep cellular structure integrated. The stability of the intermediate-type filament-lamina-nuclear matrix system in ciliates suggests that it perhaps plays an important role in ciliates cells.The protein compositions of intermediate-type filament-lamina-nuclear matrix system change after the formation of resting cyst in Urostyla grandis: there are 29 protein bands of the system in the vegetative cell and 24 protein bands in resting cyst, the former and the latter have 4 and 1 specific bands respectively, both have 23 similar bands. Concerning as the protein compositions, almost protein compositions of intermediate-type filament-lamina-nuclear matrix system i更多还原