tRNA羧甲基修饰酶Elongator在肺腺癌有丝分裂中的作用与分子机制研究

tRNA is overexpressed in cells with redundancy, and constitutes the basic module for protein translation. However, it has been recently discovered that the overexpression of some specific tRNA can drive cancer progression, meanwhile the less-expression of tRNA-modification enzyme can regulate tumorigenicity, or make tumor less metastatic. The applicant has joined one project related to the role of carboxymethyl introducing tRNA-modification enzyme Elongator in breast cancer metastasis. In this project, those codon whose decode highly depends upon Elongator, was first analyzed. The bioinformatics analysis and preliminary results indicate that Elongator can regulate cell cycle progression, and mitotic genes’ expression. Those mitotic genes regulated by Elongator, are overexpressed in lung adenocarcinoma, and those genes’ overexpression shows remarkable negative correlation with the survival of lung adenocarcinoma patients. Accordingly, this project is planned to use lung adenocarcinoma cell line, transgenic mouse model, and clinical samples, to do in-depth research upon the role of Elongator in mitosis, its molecular mechanism involved, and its translational meaning. All the work will help to understand how Elongator catalyzed tRNA modification regulates lung adenocarcinoma at mitotic level, and will also give rise to new potential targets for cancer target therapy.

tRNA作为蛋白质合成基础元件过量地存于细胞内（冗余性）。新近研究显示，改变某一种tRNA或tRNA修饰酶水平，可影响肿瘤发生、转移，提示tRNA在肿瘤发生、恶性进展过程中有一定的调控作用。申请人前期参与了tRNA羧甲基修饰酶Elongator与乳腺癌转移相关性的研究。本项目申请旨在进一步揭示Elongator参与的肿瘤生物学作用与机制，探索其靶标意义。前期功能相关密码子使用分析与实验筛查结果表明，Elongator参与调控细胞周期进行、有丝分裂蛋白表达，TCGA数据分析显示受Elongator调控的有丝分裂蛋白高表达于肺腺癌，与患者预后呈显著负相关。进而，本研究拟在细胞、转基因小鼠和临床标本水平系统阐明Elongator对肺腺癌有丝分裂的调控作用与分子机制，确立本课题相关分子的肿瘤标志物意义，揭示Elongator酶促tRNA修饰参与的肺腺癌有丝分裂调控机制，为肺腺癌治疗提供新潜在靶标。

传统理念中tRNA作为蛋白质合成基础元件，冗余、过量地存在于细胞内。新近研究揭示，tRNA，其来源新型小RNA，tiRNA与tRF，以及tRNA修饰催化酶在细胞生命活动中发挥着一定的调控性作用。tRNAGlu(UUC), tRNAGln(UUG), tRNALys(UUU)，它们的转录后成熟过程中，第34位摇摆碱基（wobble base）尿苷在羧甲基修饰酶Elongator以及硫化修饰酶CTU的作用下引入羧甲基、巯基。本项目研究解析了tRNA化学修饰酶Elongator以及CTU对有丝分裂蛋白表达的调控作用与分子机制。首先，人类基因组开放阅读框ORF的密码子分析显示密码子GAA、CAA、AAA相对富集于有丝分裂基因簇；进一步实验研究显示敲除Elongator或者CTU可以降低细胞内NUF2、HEK1、KIF4A、KIF14等有丝分裂基因的蛋白水平、增加它们mRNA上的核糖体聚集、以及它们在胞内异常蛋白聚集体中的含量，提示缺失tRNA修饰酶能够引起有丝分裂蛋白异常聚集、下调其胞内水平；蛋白质氨基酸序列分析显示，受tRNA修饰酶调控的上述有丝分裂蛋白含有共性的亲水性氨基酸短肽基序，[EKR]-[EKR]-[EKR]-R-[DEKR]，通过基因修饰突变上述氨基酸基序能够逆转缺失tRNA修饰酶所引起的有丝分裂蛋白下调。综上，本项目研究结果表明tRNA修饰能够重编程基因表达，GAA、CAA、AAA密码子富集性分布以及亲水性氨基酸序列[EKR]-[EKR]-[EKR]-R-[DEKR]的存在，与tRNA修饰酶水平共同决定蛋白质命运，调控胞内有丝分裂蛋白表达，进而调控相关生物学过程。本研究发现更新了有丝分裂调控机制的现有认识，为肿瘤靶向治疗策略的开发，提供了包括Elongator、CTU在内的tRNA修饰酶这一新型潜在靶标。