Guide RNA requirement for editing-site-specific endonucleolytic cleavage of preedited mRNA by mitochondrial ribonucleoprotein particles in Trypanosoma brucei.
Guide RNA requirement for editing-site-specific endonucleolytic cleavage of preedited mRNA by mitochondrial ribonucleoprotein particles in Trypanosoma brucei.
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布氏锥虫线粒体核糖核蛋白颗粒对预编辑 mRNA 进行编辑位点特异性核酸内切切割的指导 RNA 要求。
DOI：
10.1128/mcb.17.9.5377
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发表时间：
1997
影响因子：
5.3
通讯作者：
Hajduk,SL
中科院分区：
文献类型：
--
作者：
Adler,BK;Hajduk,SL
锥虫线粒体中的RNA编辑需要前体mrna的转录后内部添加和偶尔删除尿苷。有充分的证据表明，指定添加或删除尿苷的位置和数量的信息来自线粒体编码的小向导rna （gRNAs）。最近的研究表明，这一过程涉及酶级联反应，在编辑位点开始前mrna的核内裂解。被切割的编辑位点可以分别通过线粒体末端尿苷基转移酶（TUTase）或末端尿苷基核酸外切酶在5 ‘片段的3 ’端进行尿苷(U)的添加或删除。线粒体RNA连接酶随后重新加入mRNA。在锥虫线粒体中发现了执行这些过程的活性，包括编辑位点特异性内切酶活性，该酶可切割预编辑而非编辑的mrna。我们发现这种酶活性与先前发现的含有TUTase、RNA连接酶和grna的相同的19S核糖核蛋白颗粒共同沉积，并在盐处理后保持稳定。通过在体外添加互补寡核苷酸来消耗内源性细胞色素mrna，完全抑制合成的预编辑细胞色素mrna的编辑位点切割。添加合成的同源gRNA用于细胞染色质，但不是无关的小RNA恢复编辑位点的切割。这些研究表明，除了指定添加或删除尿苷的位点和数量外，grna还为编辑位点特异性内切酶的切割提供必要的信息。
RNA editing in trypanosome mitochondria entails the posttranscriptional internal addition and occasional deletion of uridines from precursor mRNAs. Ample evidence exists to show that the information specifying the site and number of uridines added or deleted comes from small, mitochondrially encoded guide RNAs (gRNAs). More recent work indicates that the process involves an enzymatic cascade, initiating with an endonucleolytic cleavage of the pre-mRNA at an editing site. The cleaved editing site can undergo uridine (U) addition to or deletion from the 3′ end of the 5′ fragment via a mitochondrial terminal uridylyl transferase (TUTase) or terminal uridylyl exonuclease, respectively. Mitochondrial RNA ligase subsequently rejoins the mRNA. Activities to carry out these processes have been found in trypanosome mitochondria, including an editing-site-specific endonuclease activity which cleaves preedited but not edited mRNAs. We have found that this enzymatic activity cosediments with the same 19S ribonucleoprotein particle previously shown to contain TUTase, RNA ligase, and gRNAs and remains stable after salt treatment. Depletion of endogenous cytochromebgRNAs by the addition of complementary oligonucleotides in vitro completely inhibits editing-site cleavage of synthetic preedited cytochromebmRNA. The addition of synthetic cognate gRNA for cytochromebbut not unrelated small RNA restores editing-site cleavage. These studies show that in addition to specifying the site and number of uridines added or deleted, gRNAs provide the necessary information for cleavage by the editing-site-specific endonuclease.