Novel TUTase associates with an editosome-like complex in mitochondria of Trypanosoma brucei
Novel TUTase associates with an editosome-like complex in mitochondria of Trypanosoma brucei
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DOI：
10.1261/rna.1538809
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发表时间：
2009-07-01
期刊：
影响因子：
4.5
通讯作者：
Aphasizhev, Ruslan
中科院分区：
文献类型：
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作者：
Aphasizheva, Inna;Ringpis, Gene-Errol;Aphasizhev, Ruslan
动质体原生生物线粒体基因组的表达需要大量的尿嘧啶插入/缺失mRNA编辑。级联的编辑反应由多蛋白复合物20 S编辑体完成，并由反式作用引导RNA指导。两种不同的RNA末端尿苷酰转移酶（TUTase），RNA编辑TUTase 1（RET 1）和RNA编辑TUTase 2（RET 2），分别催化向导RNA的3'尿苷酰化和向mRNA中的U-插入。RET 1也参与线粒体mRNA的转换，并参与许多异质复合物; RET 2是20 S编辑体的组成部分，其中它与锌指蛋白MP81和RNA编辑连接酶P482形成U-插入亚复合物。在这里，我们报告的第三个线粒体TUTase从布氏锥虫的鉴定。线粒体编辑体样复合物相关的TUTase（MEAT 1）与20 S编辑体样颗粒相互作用，有效地取代U-插入子复合物。MEAT 1和RET 2在它们各自的复合物中是互斥的，否则它们共享几个组件。与RET 2类似，MEAT 1在体外仅具有U特异性，并且对类似编辑底物的有缺口的双链RNA具有活性。然而，MEAT 1不需要通过编辑核酸内切酶产生的3' mRNA切割片段上的5'磷酸基团。功能性RNAi互补实验表明，MEAT 1对血流和昆虫寄生虫形式的生存力至关重要。后者中的生长抑制表型可以通过共表达RNAi抗性基因与靶向内源转录物的双链RNA来挽救。然而，初步的RNA分析显示，在MEAT 1缺失的细胞中，对RNA编辑没有明显的影响，并表明其可能在调节线粒体RNA稳定性中发挥作用。
Expression of mitochondrial genomes in Kinetoplastida protists requires massive uracil insertion/deletion mRNA editing. The cascade of editing reactions is accomplished by a multiprotein complex, the 20S editosome, and is directed by trans-acting guide RNAs. Two distinct RNA terminal uridylyl transferases (TUTases), RNA Editing TUTase 1 (RET1) and RNA Editing TUTase 2 (RET2), catalyze 3' uridylylation of guide RNAs and U-insertions into the mRNAs, respectively. RET1 is also involved in mitochondrial mRNA turnover and participates in numerous heterogeneous complexes; RET2 is an integral part of the 20S editosome, in which it forms a U-insertion subcomplex with zinc finger protein MP81 and RNA editing ligase REL2. Here we report the identification of a third mitochondrial TUTase from Trypanosoma brucei. The mitochondrial editosome-like complex associated TUTase (MEAT1) interacts with a 20S editosome-like particle, effectively substituting the U-insertion subcomplex. MEAT1 and RET2 are mutually exclusive in their respective complexes, which otherwise share several components. Similarly to RET2, MEAT1 is exclusively U-specific in vitro and is active on gapped double-stranded RNA resembling editing substrates. However, MEAT1 does not require a 5' phosphate group on the 3' mRNA cleavage fragment produced by editing endonucleases. The functional RNAi complementation experiments showed that MEAT1 is essential for viability of bloodstream and insect parasite forms. The growth inhibition phenotype in the latter can be rescued by coexpressing an RNAi-resistant gene with double-stranded RNA targeting the endogenous transcript. However, preliminary RNA analysis revealed no gross effects on RNA editing in MEAT1-depleted cells and indicated its possible role in regulating the mitochondrial RNA stability.