探討水稻OsRH2 DEAD-box RNA解旋酶之功能

Characterization of DEAD-box RNA helicase genes, OsRH2, in Oryza sativa L.

謝易軒(Yi-syuan Sie)

指導教授 ： 陸重安

國立中央大學/理學院/生命科學系/碩士(2015年)

摘要

核糖核酸解旋酶是一種機動蛋白，以三磷酸腺苷水解來產生能量將雙股核糖核酸解開的酵素。DEAD-box核糖核酸解旋酶依照其N端和C端所延伸的氨基酸具有和蛋白質結合的功能，藉著與蛋白質結合的特性，決定其受質特異性、在細胞中的位置和參與生理的功能，因此在基礎細胞中的各個過程裡包括核醣體生成、信使核糖核酸接合、核糖核酸運輸、核糖核酸轉譯和核糖核酸降解等等中扮演著相當重要的角色。目前的研究發現水稻中有62個不同的DEAD-box 核糖核酸解旋酶。此論文是研究水稻中DEAD-box 核糖核酸解旋酶，OsRH2。OsRH2為真核起始因子4AIII蛋白質，與OsRH34同為真核起始因子4AIII蛋白質且與OsRH2為高相似度蛋白。OsRH2是持續表現的一個基因且先前的研究中發現受到冷逆境所誘導。我們利用核糖核酸干擾的方式來產生轉植株並做其生理分析，經由分析我們現已獲得三個獨立植株分別命名為OsRH2Ri -2b, -4及-14b。我們從這些植株的性狀中觀察到植株矮化、較短的穗長和節間、繁殖期延後、胚胎缺失、不正常的花粉和低捻實率。這些性狀類似於吉貝素相關的賀爾蒙發育缺失，在吉貝素生合成或吉貝素感應相關蛋白質的突變株上可以觀察到。此外，OsRH2核糖核酸干擾轉植株顯示在吉貝素20氧化酶2酵素表現中發現未切除乾淨的內含子片段的信使核糖核酸有所累積。從目前的研究中我們可以發現OsRH2參與在信使核糖核酸的接合外同時影響水稻的生長發育。

關鍵字

Oryza sativa ； DEAD-box RNA helicase ； dwarf ； gibberellin

並列摘要

RNA helicases are motor proteins that catalyze the unwinding of duplex RNA/RNA in an ATP-dependent manner and thereby play important roles in most of the basic cellular processes, including ribosome biogenesis, mRNA splicing, RNA export, mRNA translation, and RNA decay. Rice (Oryza sativa) genome contains at least 62 different genes for DEAD-box RNA helicase proteins. This study investigated two DEAD box RNA helicase, OsRH2 and OsRH34, in rice. Amino acid sequence analysis indicated that the OsRH2 and OsRH34 shared 99% identity and 100% similarity. From the BiFC results, OsRH2 exhibited physical interaction with OsMAGOa and OsY14a, indicated that OsRH2 is core components of the EJC in rice. The OsRH2 were constitutively expression and activated by cold stress. RNA interference transgenic plants were generated for studying biological roles of OsRH2 and OsRH34 in rice. The phenotypes of three independent OsRH2 RNAi transgenic lines, OsRH2 RNAi -2b, -4 and -14b, indicated dwarf, short length of internode, reproduction stage delaying, embryonic development defect, abnormal pollen development, and low seed setting rate. These phenotypes resembled gibberellin-related developmental defects which were exhibited in several GA biosynthesis and GA signaling protein mutants. Besides, the OsRH2 RNAi transgenic lines exhibited unsplicing GA20ox2 mRNA accumulation. My study showed that OsRH2 and OsRH34 participate in mRNA splicing and are also involved in plant development in rice.

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