【作者】 杨帆；

【导师】 王宗华；

【作者基本信息】 福建农林大学 ， 微生物学， 2010， 硕士

【摘要】 小G蛋白是真核生物细胞信号转导过程中重要的蛋白超家族,可分为8个不同的家族:Arf、Ran、Rab、Rho、Ras、Sar1、Mitochondrial Rho、Roc small GTPases domain,拥有一个共同的结构保守的GTP结合结构域。Rho家族GTP酶主要调控细胞骨架重组。真菌中,Rho1主要调控细胞的形态建成,Rho2、Rho3以及Rho4对真菌的细胞分化和极性生长有影响。本研究通过生物信息学的方法,利用相应的计算机软件及数据库,对稻瘟病菌Rho2蛋白氨基酸序列进行分析和预测。对其理化性质、蛋白结构域、同源性、蛋白基序(motif)搜索和酶切位点预测以及蛋白序列系统演化关系做了详细的分析。进一步通过构建基因敲除突变体研究基因功能。MgRho2基因敲除突变体产孢量正常,菌落生长与野生型相比无明显差异,分生孢子和菌丝隔膜无变化,但是分生孢子的附着胞形成相对滞后于野生型,而随着时间的延长,分生孢子附着胞的形成与野生型达到相同的水平。洋葱表皮侵染实验表明,MgRho2基因敲除突变体能够产生正常的侵入栓侵入洋葱表皮,但是相对于野生型,侵入栓形成率较低,这与分生孢子的附着胞形成延迟相对应。将敲除突变体接种水稻CO39,及对大麦叶片进行离体接种,结果表明突变体发病情况与野生型没有差别。说明了不象其他Rho族成员在稻瘟病菌致病性中起重要调控作用,MgRho2主要影响该菌的附着胞发育,其机制有待进一步研究。更多还原

【Abstract】 In recent years small G proteins have become an intensively studied group of regulatory GTP hydrolases involved in cell signaling of eukaryotes. Small GTPases form an independent superfamily. This superfamily contains proteins that control a vast number of important processes and possess a common, structurally preserved GTP-binding domain. The small GTPase superfamily can be devided into eight different families: Arf, Ran, Rab, Rho, Ras, Sar1, Mitochondrial Rho, Roc small GTPases domain. Rho GTPases control cytoskeleton reorganization. In fungi, Rho1 is shown to be involved in the regulation of cell morphogenesis, Rho2, Rho3 and Rho4 are implicated in the regulation of cell differentiation and polarity growth. There exists sophisticated Rho GTPase signal pathways in the fungus, and they may regulate each other among these Rho GTPases.In this study, the amino acid sequences of Rho2 homolog in Magnaporthe oryzae was analyzed by bioinformatics and a detailed analysis of physicochemical properties, domains, homologs, motifs, enzymatic digestion sites, phylogenetic relationships are presented. Further functional analysis of Rho2 in Magnaporthe oryzae was analyzed by gene knock-out techniques. Deletion of MgRho2 had no effects on conidiation and colony growth. And the septation of conidia and hyphal were normal. But appressorium formation of MgRho2 deletion mutants were relatively delayed in spite of returning to the level of wild-type strain after further incubation. Furthermore, MgRho2 deletion mutant could penetrate onion as wild-type strain. But the rate of penetration of reduced which corresponded to delay of appressorium formation. When inoculating on barley leaves and rice leaves, there was no difference on pathogenicity between the deletion mutant and wild-type strain. These results suggest that Rho2 is not as important as other Rho members in the fungal pathogenesis, but it may regulate appressorium development of the fungus, the mechanism under which needs to further study.更多还原