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绿色植物PTP基因家族的系统进化分析及水稻OsPTP-22基因的功能研究
中文摘要

酪氨酸蛋白磷酸酶(PTP)通过去磷酸化作用,与酪氨酸激酶共同调节和维持着生物体内的酪氨酸磷酸化的适量和平衡。PTP在生物体内是一类超家族,根据催化模式的不同,PTP分为Cys-based PTP和Asp-based PTP两个类型。大量的实验证据说明PTP在对生物和非生物胁迫反应及生长和发育中起着非常重要的作用。本研究从拟南芥和水稻PTP基因,绿色植物PTP基因和OsPTP-22基因三个层次,结合物种和PTP亚家族两个角度对PTP进行了进化推理和功能研究。取得如下结果: (1)在拟南芥和水稻这两个重要模式植物中,通过全基因组分析,分别鉴定到29和39个PTP基因。对这68个PTP基因进行基因结构分析、蛋白结构域组成分析和系统进化分析。结果发现PTP基因在亚家族间高度分化而在亚家族内部高度相似,这提示PTP基因可能以亚家族为单位进化。通过染色体定位以及系统发育分析发现拟南芥PTP基因是通过区段重复方式扩增,而水稻PTP基因主要通过串联重复和区段重复扩增产生。网络表达数据分析发现拟南芥和水稻的PTP基因在多种组织/器官中具有不同的表达模式并且响应多种逆境因子,为后续功能基因组的研究提供基础。 (2)在已测序完成的主要绿色植物的4个谱系共9个物种的绿藻(Ostreococcus tauri,Ostreococcus lucimarinus,莱茵衣藻和团藻),苔藓(小立碗藓)、单子叶植物(水稻和玉米)和双子叶植物(拟南芥和白杨木)中共鉴定到248个PTP基因。根据PTP的催化模式,核心催化结构域系统发育分析和蛋白结构域组成将绿色植物中的PTP分为5类共11个亚家族。通过比较生物学的方法,发现同一绿色植物谱系物种在PTP基因数目,PTP亚家族分布和PTP蛋白结构域组成方式,而不同绿色植物谱系的物种差异则较大。 (3)基于绿色植物系统进化和结构域组成分析,推断PTP在绿色植物中可能存在以亚家族为单位逐渐增加的进化模型:在Ostreococcus tauri和Ostreococcus lucimarinus中有一个含4种亚家族的最小PTP集合“minimum PTP kit”,随着绿色植物的发展,出现两次PTP亚家族增加的现象:第一次PTP亚家族增加,可能有利于增加PTP底物类型的特异性,促进底物水解的高效率;第二次PTP亚家族增加,可能有利于植物登陆,适应疏水的陆生环境。 (4)对绿色植物和人类PTPome在基因数目,PTP亚家族和PTP结构域组成等多个方面分析发现两者之间存在巨大的差异,这提示PTPome在动植物中具有两条不同的进化路线。 (5)PTP基因亚家族中,有三种脂质磷酸酶(PTEN,MTMR,PFA-DSP)在水生绿色植物中不存在,只存在于陆生绿色植物中。这种明显的水陆植物分界的特性,提示脂质磷酸酶可能与陆生植物的登陆或登陆后的适应相关。 (6)PFA-DSP亚家族是一类脂质磷酸酶,不存在于水生植物中,只存在于陆生植物中,而且是唯一一类在人类中不存在而在陆生植物中存在的PTP亚家族。进一步发现该亚家族在单子叶植物PTP的脂质磷酸酶中数量最多。 (7)水稻PFA-DSP亚家族中的OsPTP-22基因是在水稻表达数据库中响应盐逆境最高的水稻PTP基因。体外生化活性分析结果表明OsPTP-22具有磷酸酶活性;逆境诱导实验结果表明在盐胁迫下OsPTP-22基因的转录本水平升高;酵母突变体互补实验结果表明OsPTP-22可以提高突变体对盐的耐受性。这些结果提示OsPTP-22基因是一个参与水稻对盐逆境响应的蛋白磷酸酶。 关键词:绿色植物、酪氨酸蛋白磷酸酶、水稻、脂质磷酸酶

英文摘要

Protein tyrosine phosphatases (PTP) control the balance of the protein phosphorylation in coordination with the protein tyrosine kinase (PTK). PTP is a superfamily and classified into Cys-based PTP and Asp-based PTP, according to the catalysis model. Many experimental evidences suggest that the PTP play significant roles in responses to the biotic or abiotic stresses, and organism development. We used the comparative genome methods to analysis the green plant (including the Arabidopsis thaliana and Otyza sativa) PTP genes in the angle of the species and subfamilies to find out the PTP evolution. Based on the bioinformatic analysis, we aslo adopted molecualr biology method to study the OsPTP-22 gene fucntion in the rice. The results are as follows: 1)In this study, we identified 29 and 39 PTP genes in the Arabidopsis thaliana and Oryza sativa by searching the whole genome. Based on the gene structure analysis and phylogenetic analysis of the 68 PTP genes, we found PTP were similar in the subfamily and showed great difference among the subfamilies, which suggested PTP gene may evolve in the unit of the subfamily. Based on the choromosome location and duplicaition analysis, we found that the Arabidopsis thaliana PTP genes were expanded by the segmental duplication, but the rice PTP genes were expanded by the tamdem duplication and segmental duplication. Internet expression database analysis indicated that Arabidopsis thaliana and Oryza sativa showed divese expression pattern in the tissue/organs and the response to the stress, which supplied the basis for the function analysis of the PTP genes. 2)Total 248 green plant PTP genes were identified from the four lineages including nine representative species, and were distributed among the eleven subfamilies based on the catalysis method and phylogenetic analysis of the catalytic domains amino acid sequences, which is also supported by the domain organization analysis. We also found the identical lineage species show the similar PTPome in the field of the PTP number, PTP subfamilies and PTP domain organization, with the great difference among the lineages. 3)Based on the phylogenetic analysis and domain organization, we inferred there was a PTP subfamilies stepwise increasing model following the green plant evolutionary radiation. There was a "minimum PTP kit" for tyrosine dephosphorylation in the Ostreococcus tauri and Ostreococcus lucimarinus. The first PTP subfamilies increase benefited PTP substrate specificity and functional effectiveness. The second PTP subfamilies increase provided the green plants the capacity to dephosphorylate phosphoinositide, to favor in a hydrophobic environment. 4)We found great difference between the green plant PTPome and the human PTPome, in the fields of the PTP gene number, PTP subfamilies and domain organization, which suggested the natural selection favored two different evolutionary routes in plant and animal PTP evolution. 5)Three kinds of PTP subfamilies (PTEN, MTMR and PFA-DSP), which could dephosphorylate the phosphoinositides, termed as lipid phosphatase, could not be identified in the aquatic green plants, but existed in the land green plants. The obvious difference distribution in the aquatic and land green plant indicated that the lipid phosphatases were related to the plant landing events. 6)PFA-DSP is a PTP subfamily that can dephosphorylate the phosphoinositides. Interestingly, PFA-DSP is the only subfamily that does not exit in the human PTPome. And the PFA-DSP genes are the majority lipid phosphatases in the monocots. 7)The RED data showed that the OsPTP-22 gene responsed highly to the salt stress in rice. The result of the expression pattern, yeast complementation analysis and transgenic plant analysis confirmed that the OsPTP-22 participate in response to the salt in rice. Keywords: green plant, protein tyrosine phosphatase, rice, lipid phosphatase

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