Ribosome rescue and translation termination at non-standard stop codons by ICT1 in mammalian mitochondria.
Ribosome rescue and translation termination at non-standard stop codons by ICT1 in mammalian mitochondria.
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DOI：
10.1371/journal.pgen.1004616
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发表时间：
2014-09
期刊：
影响因子：
4.5
通讯作者：
Takeuchi N
中科院分区：
文献类型：
--
作者：
Akabane S;Ueda T;Nierhaus KH;Takeuchi N
释放因子 (RF) 控制蛋白质合成的终止阶段。人类线粒体具有 1 类 RF 家族的四个不同成员：RF1Lmt/mtRF1a、RF1mt、C12orf65 和 ICT1。必需ICT1因子的同系物广泛分布于细菌和细胞器中，并且在人类线粒体中具有作为大亚基的核糖体蛋白成为核糖体一部分的奇特特征。该因子被认为可以以独立于密码子的方式拯救停滞的核糖体。该因子的作用机制尚不清楚，在此予以阐述。使用纯化组分的同源线粒体系统，我们证明集成的 ICT1 没有救援活性。相反，纯化的ICT1除了整合的拷贝外，还以化学计量与线粒体核糖体结合，并起到一般救援因子的作用，即，它从停滞在mRNA末端或中间的核糖体中，甚至从非编程核糖体中释放来自肽基tRNA的多肽。数据表明，氧化磷酸化酶 CO1 和 ND6 的有义密码子 (AGA/G) 处的异常终止也是由 ICT1 完成的，挑战了先前的模型，根据该模型，RF1Lmt/mtRF1a 负责非标准终止密码子处的翻译终止。我们还通过突变分析证明，ICT1 N 端结构域中存在的独特插入序列对于肽释放而不是核糖体结合至关重要。 RF1mt（哺乳动物线粒体中 1 类 RF 的另一个成员）的功能也进行了检查和讨论。有趣的是，哺乳动物线粒体 ICT1 是细菌 ArfB 同源物，是线粒体糖体 (MRPL58) 的组成部分。该因子的核糖体救援机制尚不清楚，在此进行讨论。利用纯化成分的同源线粒体系统，我们证明集成的 ICT1 与之前的模型相比没有救援活性。相反，添加到线粒体中的纯化ICT1具有一般的救援活性；它回收位于 mRNA 末端或中间的核糖体，甚至可以水解与非编程核糖体结合的肽基-tRNA。这些结果进一步表明ICT1可以在哺乳动物线粒体中非标准终止密码子AGA/G处的翻译终止中发挥作用。我们的数据挑战了之前的模型，该模型声称 RF1Lmt/mtRF1a 负责非标准终止密码子处的翻译终止。突变研究表明 ICT1 中的独特插入序列对于肽释放至关重要。 RF1mt（哺乳动物线粒体中 1 类 RF 的另一个成员）的功能也进行了检查和讨论。
Release factors (RFs) govern the termination phase of protein synthesis. Human mitochondria harbor four different members of the class 1 RF family: RF1Lmt/mtRF1a, RF1mt, C12orf65 and ICT1. The homolog of the essential ICT1 factor is widely distributed in bacteria and organelles and has the peculiar feature in human mitochondria to be part of the ribosome as a ribosomal protein of the large subunit. The factor has been suggested to rescue stalled ribosomes in a codon-independent manner. The mechanism of action of this factor was obscure and is addressed here. Using a homologous mitochondria system of purified components, we demonstrate that the integrated ICT1 has no rescue activity. Rather, purified ICT1 binds stoichiometrically to mitochondrial ribosomes in addition to the integrated copy and functions as a general rescue factor, i.e. it releases the polypeptide from the peptidyl tRNA from ribosomes stalled at the end or in the middle of an mRNA or even from non-programmed ribosomes. The data suggest that the unusual termination at a sense codon (AGA/G) of the oxidative-phosphorylation enzymes CO1 and ND6 is also performed by ICT1 challenging a previous model, according to which RF1Lmt/mtRF1a is responsible for the translation termination at non-standard stop codons. We also demonstrate by mutational analyses that the unique insertion sequence present in the N-terminal domain of ICT1 is essential for peptide release rather than for ribosome binding. The function of RF1mt, another member of the class1 RFs in mammalian mitochondria, was also examined and is discussed. Mammalian mitochondrial ICT1, a bacterial ArfB homolog, is interestingly an integral component of the mitoribosome (MRPL58). The mechanism of ribosome rescue by this factor was obscure and is addressed here. Utilizing a homologous mitochondria system of purified components we demonstrate that the integrated ICT1 has no rescue activity, as opposed to a previous model. Rather, purified ICT1 added to mitoribosomes has a general rescue activity; it recycles ribosomes stalled at the end or in the middle of mRNAs and can even hydrolyze peptidyl-tRNA bound to non-programmed ribosomes. These results further imply that ICT1 can function in the translation termination at non-standard stop codons AGA/G in mammalian mitochondria. Our data challenge a previous model claiming that RF1Lmt/mtRF1a is responsible for the translation termination at non-standard stop codons. A mutational study indicates that the unique insertion sequence in ICT1 is essential for peptide release. The function of RF1mt, another member of the class1 RFs in mammalian mitochondria, was also examined and is discussed.
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影响因子：
16
作者：
Tsuboi, Masafumi;Morita, Hiroyuki;Takeuchi, Nono
通讯作者：
Takeuchi, Nono
影响因子：
11.4
作者：
Richter, Ricarda;Rorbach, Joanna;Pajak, Aleksandra;Smith, Paul M.;Wessels, Hans J.;Huynen, Martijn A.;Smeitink, Jan A.;Lightowlers, Robert N.;Chrzanowska-Lightowlers, Zofia M.
通讯作者：
Chrzanowska-Lightowlers, Zofia M.
DOI：
10.1126/science.1217039
发表时间：
2012-03-16
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Neubauer C;Gillet R;Kelley AC;Ramakrishnan V
通讯作者：
Ramakrishnan V
影响因子：
64.8
作者：
Laurberg, Martin;Asahara, Haruichi;Noller, Harry F.
通讯作者：
Noller, Harry F.
影响因子：
64.8
作者：
Greber, Basil J.;Boehringer, Daniel;Ban, Nenad
通讯作者：
Ban, Nenad