I. 水稻及阿拉伯芥RNA解旋酶36之生物性功能研究 II. 阿拉伯芥中受冷誘導表現解旋酶之功能研究

I. Study of biological functions of RNA helicase 36 in Arabidopsis and rice II. Functional study of cold induced RNA helicases in Arabidopsis

黃俊凱(Chun-Kai Huang)

指導教授 ： 陸重安

國立中央大學/理學院/生命科學系/博士(2011年)

摘要

DEAD- box蛋白質參與pre-mRNA剪裁、rRNA的生合成、RNA降解及參與調控基因表現。本研究在阿拉伯芥和水稻中發現一個基因RH36，RH36可轉譯出DEAD-box蛋白質。RH36在水稻及阿拉伯芥的所有組織和器官中均有表現，經由螢光顯微鏡觀測AtRH36-GFP和OsRH36-GFP主要表現於細胞核與核仁中。在水稻及阿拉伯芥中rh36同型核子突變株為致死的表型，並且從異型核子自交的子代中異型核子植株的個數與野生型植株的個數比例為1：1，推測RH36基因可能參與在配子體的發育，藉由正反交的結果顯示AtRH36會影響雌配子體之發育。經由共軛焦顯微鏡的觀察發現雌配子體的發育會受到延遲，進而影響雌配子體之授粉。降低AtRH36基因之表現會造成植物發育上之嚴重缺失並且會大量累積pre-rRNA。經由演化樹分析比對，AtRH36與OsRH36和酵母菌的DBP8p最為相似，進一步的實驗中證明OsRH36可取代AtRH36在阿拉伯芥中的功能，但卻無法取代DBP8p在酵母菌中之功用，因此我們認為在水稻中OsRH36參與雌配子體的發育及rRNA的生合成。根據以上結果，RH36在水稻及阿拉伯芥雌配子體發育過程中之細胞分裂及rRNA的生合成中扮演著重要角色。

關鍵字

水稻 ； 阿拉伯芥 ； 解旋酶 ； 雌配子體

並列摘要

Abstract

The DEAD-box protein families are involved in RNA metabolism, including pre-mRNA splicing, ribosome biogenesis, RNA decay, and gene expression. This study indentifies a homolog of the RH36 gene encoding a DEAD-box protein in Arabidopsis, AtRH36, and rice, OsRH36. The genes were expressed ubiquitously throughout the plant. The AtRH36-GFP and OsRH36-GFP fusion protein were localized in the nucleus and nucleolus. rh36 homozygote mutants, atrh36 and osrh36, were lethal. The progenies of selfed heterozygote lines were obtained at a ratio of 1:1, as heterozygote to wild type, suggesting that the RH36 gene was involved in the gametophyte development. A reciprocal cross was therefore conducted to determine that the function of AtRH36 is via the female gametophyte. The progress of female gametogenesis was retarded in atrh36-1, and asynchronous development of female gametophyte within the same pistil was found. Knockdown of AtRH36 presented a pleiotropic phenotype and led to accumulation of unprocessed 18S pre-rRNA. In phylogenetic analysis, AtRH36 and OsRH36 were homologous to yeast DBP8p. Furthermore, functional complementation tests among three homologous DEAD-box protein genes indicated that OsRH36 can restore segregation distortion in the Arabidopsis atrh36-1 mutant, but cannot complement the lethal phenotype in the yeast dbp8p mutant. Therefore, we suggest that OsRH36 is required for rRNA biogenesis and megagametogenesis in rice, consistent with the function of AtRH36 in Arabidopsis. These results suggest that RH36 is essential for the mitotic division cycles during development of female gametogenesis and plays an important role in rRNA biogenesis in rice and Arabidopsis.

並列關鍵字

rice ； arabidopsis ； RNA helicase ； female gametophyt

參考文獻

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