CCACA碱基序列是玉米CMS-S线粒体orf355-orf77转录本剪切识别位点

Identification of Sequence CCACA as a Recognition Motif of Mitochondrial Transcripts Orf355-orf77 Cleavage Sites in CMS-S Maize

玉米S型细胞质雄性不育系(CMS-S)及其近等基因恢复系是研究核-质互作机制的重要遗传资源和理想模式体系.目前认为,CMS-S花粉败育是由其线粒体内细胞质不育基因orf355-orf77表达的毒性蛋白引起,而核育性恢复基因Rf3可通过引发orf355-orf77转录本的降解而解除其毒性作用,使花粉育性得以恢复.本研究采用Northern杂交和3'RACE技术确定了orf355-orf77转录本的剪切位点,并发现在育性恢复的花粉中,orf355-orf77转录本被剪切成小片段之后聚合了poly(A)序列,推测这一过程加速了mRNA分子的降解,是育性恢复的关键环节.利用生物信息学方法分析了orf355-orf77转录本6个剪切位点的侧翼序列,发现在剪切位点下游10个碱基的位置均含有5'-CCACA-3'序列,推测该序列受到特定功能蛋白的识别,然后募集核酸内切酶对其进行剪切.研究结果可为揭示玉米CMS-S育性恢复机理提供重要的理论依据.

S-type cytoplasmic male sterility line （CMS-S） and its near isogenic restorer lines of maize are suitable materials for the study on nucleus-mitochondria transcript interactions. The toxicity product coded by orf355-orf77 is considered to be potentially inducible in CMS-S. Its fertility restorer, Rf3 gene can induce the degradation and removal of orf355-orf77 transcripts of its toxic effect, and protect the development of microspores in normal status. In this study, the cleaving sites of orf355-orf77 transcript was determined by Northern blotting and 3 RACE. We found that most of the small cleaved fragments from accelerated RNA degradation, were polyadenylated. Through bioinformatic analysis of the six flanking sequences of cleaved sites of orf355-orf77 transcripts, a conserved recognition pattern of CCACA was identified at the 10 nucleotides downstream of the cleavage sites. This sequence was predicted to be recognizable by a specific protein to cleave the RNAs by recruiting endonuclease. The results may provide important information in revealing the mechanism of CMS-S fertility restoration in maize.