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首页> 美国卫生研究院文献>The Plant Cell >Synthesis of the D2 Protein of Photosystem II in Chlamydomonas Is Controlled by a High Molecular Mass Complex Containing the RNA Stabilization Factor Nac2 and the Translational Activator RBP40
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Synthesis of the D2 Protein of Photosystem II in Chlamydomonas Is Controlled by a High Molecular Mass Complex Containing the RNA Stabilization Factor Nac2 and the Translational Activator RBP40

机译：衣藻中光系统II的D2蛋白的合成受包含RNA稳定因子Nac2和翻译激活因子RBP40的高分子复合物控制。

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摘要

Gene expression in chloroplasts is regulated mainly at the posttranscriptional level. In the green alga Chlamydomonas reinhardtii, synthesis of the D2 protein (PsbD), which is the rate-determining subunit for the assembly of photosystem II, depends on the RNA stability factor Nac2. In addition, the RNA binding protein RBP40 has been implicated in translational control via a U-rich element in the 5′ untranslated region (5′UTR) of the psbD mRNA. Here, we report the identification of the RBP40 gene based on mass spectrometric analysis of its purified product. Unexpectedly, this was found to be identical to the previously described RNA binding protein RB38, which had been suggested to be involved in the regulation of D1 protein synthesis. However, we show that RBP40 binds to the psbD 5′UTR in a Nac2-dependent fashion both in vitro and in vivo. Molecular characterization of RBP40 RNA interference lines confirmed that RBP40 specifically affects the initiation of D2 synthesis. Native polyacrylamide gel electrophoresis, coimmunoprecipitation, and sedimentation analyses revealed that Nac2 and RBP40 form parts of a complex of 550 kD that is displaced from the psbD mRNA prior to polysome assembly. Together, these data indicate that the processes of 5′UTR-mediated RNA stabilization and translation initiation are tightly coupled in Chlamydomonas.
机译：叶绿体中的基因表达主要在转录后水平调控。在绿藻莱茵衣藻中，D2蛋白（PsbD）的合成是光系统II装配的决定速率的亚基，它的合成取决于RNA稳定因子Nac2。另外，RNA结合蛋白RBP40已经通过psbD mRNA的5'非翻译区（5'UTR）中的富含U的元件参与翻译控制。在这里，我们报告基于其纯化产物的质谱分析鉴定RBP40基因。出乎意料的是，发现它与先前描述的RNA结合蛋白RB38相同，后者已被证明与D1蛋白合成的调控有关。然而，我们显示RBP40在体外和体内均以依赖于Nac2的方式与psbD 5'UTR结合。 RBP40 RNA干扰品系的分子表征证实，RBP40特别影响D2合成的启动。天然聚丙烯酰胺凝胶电泳，免疫共沉淀和沉降分析表明，Nac2和RBP40形成550 kD复合物的一部分，该复合物在多核糖体组装前已从psbD mRNA移出。总之，这些数据表明5'UTR介导的RNA稳定和翻译起始过程在衣藻中紧密耦合。

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