【作者】 陆玉建；

【导师】 郑国锠；

【作者基本信息】 兰州大学 ， 细胞生物学， 2010， 博士

【摘要】 在高等真核生物进化的过程中,ZW10蛋白是相当保守的。大量的研究已经证实,在动物细胞中,ZW10无论对于有丝分裂还是减数分裂过程中染色体的准确分离都是十分重要的。在细胞分裂的间期,ZW10蛋白还参与内质网和高尔基体之间的膜泡运输。多重序列比对显示,不同生物的ZW10蛋白序列中存在许多一致的区域,表明在不同的有机体中,ZW10蛋白可能具有相似的功能。本文对拟南芥中ZW10同源蛋白-AtZW10-进行了初步研究。RT-PCR和GUS染色结果显示,AtZW10在不同的组织和器官中均有表达,但在花和种子中表达量更高。亚细胞定位分析表明,这一蛋白在细胞核和细胞质中均有分布,但在细胞核中分布更为显著。通过分析该基因的3个T-DNA插入突变体,结果显示,在正常的生长条件下,这些突变体的表型与野生型没有明显的差异;但在不同的胁迫条件下,突变体种子萌发比野生型要快的多,这些数据表明AtZW10很可能作为种子萌发的负调控因子参与种子萌发的过程。进一步的研究证实,AtZW10影响细胞分裂和早期胚胎发育的过程。在zw10-2突变体中,与野生型相比,有丝分裂和减数分裂都存在明显的缺陷。在突变体有丝分裂的后期,可以观察到染色体分离延迟现象；而在减数分裂的突变体细胞中,异常的分裂可出现在细胞周期的不同阶段,这些异常包括染色体片段,不均等或不同步的染色体分离,以及延迟的染色体等。并且,实验结果显示,AtZW10功能缺失将导致拟南芥胚胎发育的早期出现明显的抑制,多数突变体胚胎被抑制在2-细胞时期。这些突变体胚胎的细胞分裂面出现异常,纵向分裂常常被横向分裂所取代,并且还出现低频率的独特的分裂方式。此外,初步的分析显示,在拟南芥中,MAG2蛋白(即动物细胞中RINT-1蛋白同源物)似乎能与AtZW10蛋白相互作用,并参与内质网和高尔基体之间的膜泡运输。总之,结果表明,AtZW10是一种在进化上相当保守的蛋白,具有多种重要的生物功能。更多还原

【Abstract】 The ZW10 protein is much conserved during the evolution of higher eukaryotes. In animal cells, a large number of studies show that ZW10 is required for proper chromosome segregation during both mitosis and meiosis. The protein is also found to be involved in membrane trafficking between the endoplasmic reticulum (ER) and Golgi apparatus. Multiple sequence alignment of ZW10 homologues shows that there are a number of identical regions in ZW10 sequences, suggesting that the proteins in different organisms may have similar functions. In this paper, AtZW10, a homologous protein of ZW10, was studied in Arabidopsis. Reverse transcription-PCR analysis andβ-glucuronidase staining show that AtZW10 is expressed in different organs. However, this expression is much stronger in flowers and seeds. The subcellular localization of AtZW10 was also examined, and it was found to be localized in the nucleus and cytoplasm. However, the distribution in the nucleus even more significant. To investigate AtZW10 function, T-DNA insertion mutants were analyzed. The phenotypes of zw10 mutants were undistinguishable from those of wild-type plants under normal growth conditions. Under different stresses, mutant seeds germinated obviously faster than the wild type. These data suggested that AtZW10 may be implicated in the process of seed germination as a negative regulator.Further research showed that AtZW10 affected cell division and early embryonic development. In mitotic mutant cells, lagging chromosomes were seen. The process took place at anaphase. Additionally, many defects were found in the meiotic mutant cells, which occurred at different stages of cell division. These anomalies include chromosome fragments, unequal or asynchronous segregation of chromosomes, and lagging chromosomes.Moreover,the loss-of-function mutations of AtZW10 caused the arrest of early embryo development. Most mutant embryos were found to be arrested at the two-cell stage. In addition, the cell division pattern of the mutant embryos is often changed. Longitudinal division planes of the proembryo are frequently replaced by transverse divisions and less frequently by unique divisions.Preliminary analysis shows Arabidopsis MAG2 (RINT-1 in animal cells) also seemed to interact with AtZW10 and to be involved in the membrane trafficking between the ER and the Golgi complex. In short, results showed that AtZW10,a rather conservative protein in evolution, has a variety of important biological functions.更多还原