Processing of the 5'-UTR and existence of protein factors that regulate translation of tobacco chloroplast psbN mRNA.
Processing of the 5'-UTR and existence of protein factors that regulate translation of tobacco chloroplast psbN mRNA.
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5-UTR 的加工和调节烟草叶绿体 psbN mRNA 翻译的蛋白质因子的存在。
DOI：
10.1007/s11103-014-0248-z
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发表时间：
2014
期刊：
影响因子：
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通讯作者：
H. and M. Sugiura
中科院分区：
文献类型：
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作者：
Kuroda;H. and M. Sugiura
叶绿体基因包括5个编码光系统II和细胞色素b6/f复合体成分的基因。PsbN基因位于相反的链上。PsbN定位于类囊体中，即使在黑暗中也存在，尽管它的水平在光照后增加，然后下降。然而，psbNmRNA的翻译机制仍不清楚。利用烟草叶绿体的体外翻译系统，以绿色荧光蛋白为报告蛋白，我们证明了47个核苷酸(NT)的烟草原生体bN5‘-UTR发生了翻译。与许多其他叶绿体5‘-UTRs不同，psbN5’-UTRs有两个加工位点，分别位于起始点上游的−39和−24。在−39的处理将翻译率提高了五倍。相比之下，−24的处理不影响转换率。这些观察表明，这两个不同的加工事件至少部分地调节了发育过程中PsbN的水平。PsbN5‘-非编码区没有Shine-Dalgarno(SD)样序列。体外翻译分析表明，过量的psbN5‘-UTR或缺失的psbN5’-UTR序列是翻译所必需的蛋白质因子，并且它们的结合部位是5‘-UTR中的18个核苷酸序列。用另外10个叶绿体5‘-UTRs进行的迁移率漂移分析表明，共同或类似的蛋白质参与了一组缺乏SD类似序列的mRNAs的翻译。
The chloroplastpsbBoperon includes five genes encoding photosystem II and cytochromeb6/fcomplex components. ThepsbNgene is located on the opposite strand. PsbN is localized in the thylakoid and is present even in the dark, although its level increases upon illumination and then decreases. However, the translation mechanism of thepsbNmRNA remains unclear. Using an in vitro translation system from tobacco chloroplasts and a green fluorescent protein as a reporter protein, we show that translation occurs from a tobacco primarypsbN5′-UTR of 47 nucleotides (nt). Unlike many other chloroplast 5′-UTRs, thepsbN5′-UTR has two processing sites, at −39 and −24 upstream from the initiation site. Processing at −39 enhanced the translation rate fivefold. In contrast, processing at −24 did not affect the translation rate. These observations suggest that the two distinct processing events regulate, at least in part, the level of PsbN during development. ThepsbN5′-UTR has no Shine–Dalgarno (SD)-like sequence. In vitro translation assays with excess amounts of thepsbN5′-UTR or with deletedpsbN5′-UTR sequences demonstrated that protein factors are required for translation and that their binding site is an 18 nt sequence in the 5′-UTR. Mobility shift assays using 10 other chloroplast 5′-UTRs suggested that common or similar proteins are involved in translation of a set of mRNAs lacking SD-like sequences.