水稻胞质不育相关蛋白ORF290及互作蛋白对酵母生长的影响  

Effects of ORF290 Related with Rice Cytoplasmic Male Sterility and Its Interaction Protein on Yeast Growth

作　　者：夏栋 谭艳平 刘学群 刘新琼 李开 王春台 Xia Dong;Tan Yanping;Liu Xuequn;Liu Xinqiong;Li Kai;Wang Chuntai(Key Lab for Biotechnology of State Ethnic Affairs Commission,Hubei Provincial Key Laboratory for Protection and Application of Special Plants in Wuling Area of China,South-Central University for Nationalities,Wuhan,430074)

机构地区：[1]中南民族大学生命科学学院武陵山区特色资源植物种质保护与利用湖北省重点实验室生物技术国家民委重点实验室,武汉430074

出　　处：《分子植物育种》2023年第9期2933-2940,共8页Molecular Plant Breeding

基　　金：国家自然科学基金面上项目(31170296)资助。

摘　　要：orf290是本实验室从红莲型水稻细胞质雄性不育品种‘粤泰A’(YTA)中克隆的一个与雄性不育相关的线粒体基因,前期通过膜蛋白酵母双杂交获得了2个与ORF290互作的蛋白APX8和E37。为探讨ORF290与APX8和E37发生相互作用时对酿酒酵母NMY51生长的影响,本实验利用膜蛋白酵母双杂交DUAL membrane系统,分别将诱饵蛋白和互作蛋白基因构建到pBT3-SUC-和pPR3-N-载体上,获得pBT3-SUC-orf290、pPR3-N-APX8和p PR3-N-E37质粒,然后分别单转和共转(pBT3-SUC-orf290与pPR3-N-APX8,pBT3-SUCorf290与pPR3-N-E37)酵母NMY51菌株。在RNA和蛋白水平分别证实相关基因在转化酵母中高效表达,并且相关蛋白以膜蛋白的形式存在。含不同质粒酵母生长曲线测定结果表明,宿主菌NMY51、单独转化pBT3-SUC-orf290、p PR3-N-APX8和pPR3-N-E37的NMY51酵母生长曲线相近,体内ATP含量无明显差异,而共表达ORF290与APX8或E37的酵母菌株的生长明显低于对照组,ATP含量也明显降低。这表明ORF290与APX8或E37互作后通过降低体内ATP水平导致酵母生长受到抑制。A cytoplasmic male sterility related mitochondrial gene orf290 was cloned from HL-CMS rice YTA in the laboratory.In the early stage,we obtained two interact proteins APX8 and E37 which interact with ORF290through membrane-based yeast two-hybrid.To explore the effect of ORF290 on the growth of Saccharomyces cerevisiae NMY51 when it interacts with APX8 and E37,the membrane-based yeast two-hybrid DUAL membrane system was used to construct bait protein gene and interaction protein genes into pBT3-SUC and pPR3-N vectors in this experiment.The plasmids pBT3-SUC-orf290,pPR3-N-APX8 and pPR3-N-E37 were constructed,and transformed or co-transformed(pBT3-SUC-orf290 and pPR3-N-APX8,pBT3-SUC-orf290 and pPR3-N-E37)into yeast strain NMY51 respectively.It was confirmed at the RNA and protein levels that related genes were highly expressed in transformed yeast,and related proteins existed in the form of membrane proteins.The results show that the growth curves of the host strain NMY51 and the NMY51 yeast strains transformed with pBT3-SUC-orf290,p PR3-N-APX8 and p PR3-N-E37 are similar,and there is no significant difference in the ATP content in the cell,while the growth of yeast strains co-expressing ORF290 and APX8 or E37 was significantly slower than that of the control group,and the ATP content was also significantly reduced.This indicates that ORF290 reduces the level of ATP in the yeast cell after interacting with APX8 or E37,resulting in the inhibition of yeast growth.

关 键 词：细胞质雄性不育 线粒体 orf290 互作蛋白 酵母生长 

分 类 号：S511[农业科学—作物学]