Center for Gene Research, Nagoya University, Chikusa-ku, Nagoya 464-8602, Japan.
Plant J. 2010 May 1;62(4):560-70. doi: 10.1111/j.1365-313X.2010.04175.x. Epub 2010 Feb 16.
In most land plants RNA editing frequently occurs in many organelle transcripts, but little is known about the molecular mechanisms of the organelle RNA editing process. In this study, we have characterized the Physcomitrella patens PpPPR_71 gene that is required for RNA editing of the ccmFc transcript. This transcript harbors two RNA editing sites, ccmF-1 and ccmF-2, that are separated by 18 nucleotides. Complementary DNA sequence analysis of ccmFc suggested that RNA editing at the ccmF-1 site occurred before ccmF-2 editing. RNA editing of the ccmF-2 downstream site was specifically impaired by disruption of the PpPPR_71 gene that encodes a polypeptide with 17 pentatricopeptide repeat motifs and a C-terminal DYW domain. The recombinant PpPPR_71 protein expressed in Escherichia coli specifically bound to the 46-nucleotide sequence containing the ccmF-2 editing site. The binding affinity of the recombinant PpPPR_71 was strongest when using the edited RNA at ccmF-1. In addition, the DYW domain also binds to the surrounding ccmF-2 editing site. We conclude that PpPPR_71 is an RNA-binding protein that acts as a site recognition factor in mitochondrial RNA editing.
在大多数陆地植物中,RNA 编辑经常发生在许多细胞器转录本中,但关于细胞器 RNA 编辑过程的分子机制知之甚少。在这项研究中,我们对拟南芥 PpPPR_71 基因进行了表征,该基因是 ccmFc 转录本 RNA 编辑所必需的。该转录本含有两个 RNA 编辑位点 ccmF-1 和 ccmF-2,它们之间相隔 18 个核苷酸。ccmFc 的 cDNA 序列分析表明,ccmF-1 位点的 RNA 编辑发生在 ccmF-2 编辑之前。破坏编码具有 17 个五肽重复基序和 C 端 DYW 结构域的多肽的 PpPPR_71 基因,特异性地损害了 ccmF-2 下游位点的 RNA 编辑。在大肠杆菌中表达的重组 PpPPR_71 蛋白特异性地结合到含有 ccmF-2 编辑位点的 46 个核苷酸序列上。当使用在 ccmF-1 处编辑的 RNA 时,重组 PpPPR_71 的结合亲和力最强。此外,DYW 结构域也与周围的 ccmF-2 编辑位点结合。我们得出结论,PpPPR_71 是一种 RNA 结合蛋白,作为线粒体 RNA 编辑中的位点识别因子发挥作用。