Study on the function of RNA0binding proteins of cyanobcteria and chloroplasts

蓝藻和叶绿体RNA0结合蛋白的功能研究

• 批准号: 07454204
• 负责人: SUGITA Mamoru
• 金额: $ 4.67万
• 依托单位: Nagoya University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1995
• 资助国家: 日本
• 起止时间: 1995 至 1996
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07454204/
• 关键词: chloroplast; RNA-binding protein; cyanobacteria; ribonucleoprotein particle; pre-mRNA processing; posttranscriptional regulation; mRNAプロセシング; 遺伝子破壊; 遺伝子

1. RNA-binding proteins, Rbp1 and Rbp2, of the unicellular cyanobacterium Synechococcus sp, strain PCC6301 are localized in cytosol and bind RNA molecules in vivo. To elucidate their functions, gene-disruption experiments were performed. The experiments indicated that disruption of rbpl gene is lethal while rbp2 gene is not essential under normal growth condition. We are currently identifying the target RNA species of Rbp proteins.2. To clarify the function of chloroplast RNA-Binding proteins (cpRNPs), effect of the cpRNPs on pre-mRNA processing was assayd. When cpRNPs were removed from in vitro assay system, processing of pre-mRNA was enhanced. This suggests that cpRNPs may take part in mRNA processing as negative regulator.3. cDNA2s encoding RNA-binding proteins, RZ-1 and RGP-3, have been isolated from a Nicotiana sylvestris cDNA library. The deduced amino acid sequences revealed that RZ-1 contains a consensus sequence-type RNA-binding domain (CS-RBD) and a zinc finger motif of the CCHC type, and RGP-3 has a CS-RBD and a short glycine-rich sequence. RZ-1 is localized in the nucleoplasm of tobacco cultured cells. Glycerol gradient fractionation of tobacco nuclear lysates showed the tRZ-1 is associated with a large ribonucleoprotein particle of 60 S in size. This suggests that RZ-1 may be involved in pre-mRNA processing. RGP-3 is localized as 10S particles in nucleoplasm. The biochemical basis of the RNA specificity and of ribonucleoprotein particles is an interesting issue, and further experiments are necessary to elucidate the function of RZ-1 and RGP-3.

1. 单细胞蓝藻聚藻球菌PCC6301的RNA结合蛋白Rbp1和Rbp2定位于胞浆中，并在体内结合RNA分子。为了阐明它们的功能，我们进行了基因破坏实验。实验表明，在正常生长条件下，rbpl基因的破坏是致命的，而rbp2基因的破坏则不是必需的。我们目前正在鉴定Rbp蛋白的靶RNA种类。为了阐明叶绿体rna结合蛋白（cpRNPs）的功能，研究了cpRNPs对mrna前体加工的影响。当体外实验系统中去除cpRNPs时，pre-mRNA的加工增强。这提示cpRNPs可能作为负调控因子参与mRNA加工。从烟叶cDNA文库中分离到编码rna结合蛋白RZ-1和RGP-3的cDNA2s。推导出的氨基酸序列表明，RZ-1含有一个一致序列型rna结合域（CS-RBD）和一个CCHC型锌指基序，RGP-3含有CS-RBD和一个短的富含甘氨酸的序列。RZ-1定位于烟草培养细胞的核质中。烟草核裂解物的甘油梯度分离表明，tRZ-1与一个大的60 S大小的核糖核蛋白颗粒有关。这表明RZ-1可能参与了pre-mRNA的加工。RGP-3在核质中定位为10S粒子。RNA特异性和核糖核蛋白颗粒的生化基础是一个有趣的问题，需要进一步的实验来阐明RZ-1和RGP-3的功能。

项目成果

守口和基: "Structure and subcellular localization of a small RNA-binding protein from tobacco." Plant Journal. (印刷中). (1997)

Kazuki Moriguchi：“烟草中小 RNA 结合蛋白的结构和亚细胞定位”（正在出版）。

Fujishiro, Tsuneo: "Organization and transcription of a putative gene cluster encoding ribosomal protein S14 and an oligopeptide permease-like protein in the cyanobacterium Synechococcus PCC 6301." DNA Res.3. 165-169 (1996)

Fujishiro, Tsuneo：“蓝藻聚球藻 PCC 6301 中编码核糖体蛋白 S14 和寡肽通透酶样蛋白的假定基因簇的组织和转录。”

杉田 護: "Regulation of gene expression in chloroplasts of higher plants." Plant Molecular Biology. 32. 315-326 (1996)

Mamoru Sugita：“高等植物叶绿体中基因表达的调节。”植物分子生物学 32. 315-326 (1996)

杉田 護: "Genes encoding the group I intron-containing tRNAL eu and subunit L of NADH dehydrogenase from the cyanobacterium Synechococcus RCC 6301." DNA Research. 2. 71-76 (1995)

Mamoru Sugita：“编码来自蓝细菌聚球藻 RCC 6301 的包含 I 组内含子的 tRNAL eu 和 NADH 脱氢酶 L 亚基的基因。”2. 71-76 (1995)。

花野滋: "Isolation of a novel RNA-binding protein and its association with a large ribonucleoprotein particle present in the nucleoplasm of tobacco cells." Plant Molecular Biology. 31. 57-68 (1996)

Shigeru Hanano：“一种新型 RNA 结合蛋白的分离及其与烟草细胞核质中存在的大核糖核蛋白颗粒的关联。”植物分子生物学 31. 57-68 (1996)。