拟南芥RabD2b保守半胱氨酸的缺失或突变影响其定位和功能  

Membrane Localization and Function of Arabidopsis RabD2b was Influenced by the Lacking or Mutation of the Conserved Cystein Residues

作　　者：王芳[1] 刘超[1] 吴江生[1] 刘克德[1] 

机构地区：[1]华中农业大学植物科学技术学院,武汉430070

出　　处：《农业生物技术学报》2015年第1期12-19,共8页Journal of Agricultural Biotechnology

基　　金：国家自然科学基金项目(No.30623012)

摘　　要：Rab蛋白羧基末端含有保守的半胱氨酸残基,是蛋白质翻译后进行异戊二烯化修饰的位点。本研究以拟南芥(Arabidopsis thaliana)突变型Rab D2b为对象,用定点突变的方法,获得At Rab D2b羧基末端第199位和200位保守半胱氨酸残基共同缺失的突变基因At Rab D2bΔCC,及第199位保守半胱氨酸残基突变为丝氨酸的基因At Rab D2bC199S。将突变基因瞬时转化烟草(Nicotiana tabacum)叶片或稳定转化拟南芥,发现两种突变蛋白都定位在细胞质中,而野生型At Rab D2b蛋白则主要定位在高尔基体上。酵母温敏突变体ypt1产生的温度致死表型能够通过转化At Rab D2b获得恢复,而突变后的At Rab D2b均丧失了恢复功能。结果表明,拟南芥At Rab D2b羧基末端单个保守半胱氨酸残基的突变或两个保守半胱氨酸的缺失都会影响其亚细胞定位和功能发挥。本研究为深入开展植物Rab蛋白的异戊二烯化修饰机制提供参考资料。Rab proteins have the conserved cystein residues at C terminus, which are the prenylational modification site after protein translation. In this study, we obtained Arabidopsis RabD2b mutant alleles with either lacking both two conserved cystein residues at 199 and 200 （AtRabD2bacc） or altering one conserved cystein residue at 199 to serine residue （AtRabD2b） by site mutation. When the mutated genes were transiently transformed into tobacco（Nicotiana tabacum） leaves or stably transformed into Arabidopsis, it was found that they were both targeted to the cytoplasm, while wild-type AtRabD2b was mainly localized on Golgi stacks. The lethal phenotype at certain temperature of Yeast Yptl, a temperature-sensitive mutant stain, could be recovered by the transformation of AtRabD2b. However, two mutant AtRabD2b proteins both lost the complementary ability. These results suggested that the mutation of single conserved cystein residue or lacking of the two conserved cystein residues at C terminus affected the localization and function of AtRabD2b. The results provide the reference data for further study on the mechanism of plant Rab prenylational modification.

关 键 词：保守半胱氨酸 异戊二烯化修饰 定位 功能 

分 类 号：Q939.4[生物学—微生物学]