Kinetoplastid guide RNA biogenesis is dependent on subunits of the mitochondrial RNA binding complex 1 and mitochondrial RNA polymerase
Kinetoplastid guide RNA biogenesis is dependent on subunits of the mitochondrial RNA binding complex 1 and mitochondrial RNA polymerase
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DOI：
10.1261/rna.1411809
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发表时间：
2009-04-01
期刊：
影响因子：
4.5
通讯作者：
Lukes, Julius
中科院分区：
文献类型：
--
作者：
Hashimi, Hassan;Cicova, Zdenka;Lukes, Julius
线粒体RNA结合复合体1(MRB1)是最近在布鲁氏锥虫中发现的与TbRGG1和TbRGG2蛋白相关的蛋白质复合体。基于这两种蛋白下调所引起的表型，有人认为它在RNA编辑中扮演了一个未指明的角色。三个新鉴定的蛋白质亚基，引导RNA相关蛋白(GAP)1和2以及预测的DExD/H-box RNA解旋酶的RNAi沉默表明，它们对细胞在原周期阶段的生长是必不可少的。此外，它们的下调导致仅在需要小环编码的引导(G)RNA的那些mRNA中抑制编辑。然而，当使用Maxicircle编码的顺式作用gRNA时，编辑仍然不受影响。有趣的是，所有这三种蛋白质都是表达微环编码的gRNA所必需的。此外，第四个被检测的MRB1亚基Nudex水解酶的下调似乎不会破坏gRNAs的稳定，并且该蛋白的下调通常会对Maxicircle编码的RNA的稳定性产生影响。Gap1和Gap2对于血流阶段的生存也是必不可少的，在这一阶段，当两种蛋白质中的任何一种被耗尽时，gRNA就会被消除。免疫组织化学定位显示，Gap1和Gap2集中在线粒体上的离散斑点中，通常位于动质粒附近。最后，我们证明了相同的mtRNA聚合酶转录上环mRNAs也可能在表达小环编码的gRNAs中发挥作用。
The mitochondrial RNA binding complex 1 (MRB1) is a recently discovered complex of proteins associated with the TbRGG1 and TbRGG2 proteins in Trypanosoma brucei. Based on the phenotype caused by down-regulation of these two proteins, it was proposed to play an unspecified role in RNA editing. RNAi silencing of three newly characterized protein subunits, guide RNA associated proteins (GAPs) 1 and 2 as well as a predicted DExD/H-box RNA helicase, show they are essential for cell growth in the procyclic stage. Furthermore, their down-regulation leads to inhibition of editing in only those mRNAs for which minicircle-encoded guide (g) RNAs are required. However, editing remains unaffected when the maxicircle-encoded cis-acting gRNA is employed. Interestingly, all three proteins are necessary for the expression of the minicircle-encoded gRNAs. Moreover, down-regulation of a fourth assayed putative MRB1 subunit, Nudix hydrolase, does not appear to destabilize gRNAs, and down-regulation of this protein has a general impact on the stability of maxicircle-encoded RNAs. GAP1 and 2 are also essential for the survival of the bloodstream stage, in which the gRNAs become eliminated upon depletion of either protein. Immunolocalization revealed that GAP1 and 2 are concentrated into discrete spots along the mitochondrion, usually localized in the proximity of the kinetoplast. Finally, we demonstrate that the same mtRNA polymerase known to transcribe the maxicircle mRNAs may also have a role in expression of the minicircle-encoded gRNAs.