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Translation Recoding Control

翻译重新编码控制

基本信息

批准号：
14035246
负责人：
MATSUFUJI Senya
金额：
$ 63.1万
依托单位：
Jikei University School of Medicine
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research on Priority Areas
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2006
项目状态：
已结题

项目摘要

"Recoding" is non-standard genetic decoding such as translational frameshifting. Antizyme, a regulatory protein for polyamines, requires translational frameshifting for its expression and induction by polyamines. We have identified the signal sequences, namely, the shift site, a downstream pseudoknot, and an upstream sequence, for the frameshifting of AZ1, an major isoform of AZ. This project aimed to understand the mechanism of AZ1 frameshifting form the aspects of not only the cis-acting signals but also translational machinery and small-molecule regulators, and the results are summarized as the followings. (1) The molecular evolution, interchangeability with other recoding signals, and the interaction site of the polyamine action of the AZ1 frameshift signal were analyzed. A cellular binding protein for AZ1 pseudoknot has been identified. Some small nucleotides that complementarily bind to the frameshift signals are found to affect frameshift efficiency. (2) In the ribosomal hopping directed by the mammalian AZl sequence in E. coli, re-pairing of the peptidyl-tRNA is found to be major determinant of the landing codon. (3) To identify translation machinery involved in AZ1 frameshifting, genetic screening was performed in the fission yeast. We screened genomic mutations, multicopy suppressors, dominant suppressors, but could not obtain any significant candidate. (4) Some branched-chain polyamines originated from a thermophilic bacteria or acetylated forms of polyamines showed specific effect on AZ1 frameshifting. (5) To discover new recoding genes, we screed a random sequence pool in E. coli. We also searched mRNAs of which expression levels are elevated in the nonsense-mediated mRNA decay mutants in the fission yeast. Both screening brought about several candidate for new recoding. (6) Analysis of AZ1 knockout mice revealed that a phenotype of lack of frameshift regulation of AZl is embryonic death due to a disturbance of hematopoietic cell differentiation.

“重新编码”是非标准遗传解码，例如平移移码。抗酶是多胺的调节蛋白，需要翻译移码才能表达并被多胺诱导。我们已经确定了 AZ1（AZ 的主要亚型）移码的信号序列，即移位位点、下游假结和上游序列。本项目旨在从顺式作用信号、翻译机制和小分子调节因子等方面了解AZ1移码机制，结果总结如下。 (1)分析了AZ1移码信号的分子进化、与其他重编码信号的互换性以及多胺作用的相互作用位点。已鉴定出 AZ1 假结的细胞结合蛋白。一些与移码信号互补结合的小核苷酸被发现会影响移码效率。 (2)在大肠杆菌中由哺乳动物AZ1序列指导的核糖体跳跃中，发现肽基-tRNA的重新配对是着陆密码子的主要决定因素。 (3)为了鉴定参与AZ1移码的翻译机制，在裂殖酵母中进行了遗传筛选。我们筛选了基因组突变、多拷贝抑制基因、显性抑制基因，但未能获得任何重要的候选基因。 (4)一些源自嗜热细菌的支链多胺或多胺的乙酰化形式对AZ1移码显示出特定的作用。 (5) 为了发现新的重新编码基因，我们在大肠杆菌中建立了一个随机序列库。我们还在裂殖酵母中的无义介导的 mRNA 衰变突变体中搜索了表达水平升高的 mRNA。两次筛选都带来了一些新编码的候选者。 (6)对AZ1敲除小鼠的分析表明，缺乏AZ1移码调节的表型是由于造血细胞分化紊乱而导致的胚胎死亡。