| 摘要: | 基因表現的過程中，RNA分子(mRNA, rRNA, and tRNA)扮演許多角色。RNA分子為單股，在環境逆境易形成不正常的構型。DEAD-box RNA helicases為細胞中維持或修改不同標的RNA分子正確構型的重要蛋白質之一。因此，當植物細胞處於環境逆境時，DEAD-box RNA helicases能維持RNA分子的正常功能，是細胞維持正常基因表現與逆境耐受力的重要關鍵之一。水稻為世界上重要糧食作物，是熱帶和亞熱帶作物，對低溫相當敏感。水稻至少含有62種不同的基因編碼DEAD-boxRNA helicase，不同DEAD box RNA helicase在生物體內有特殊生理功能。先前，我們發現OsRH42為受冷逆境誘導的基因，OsRH42直接結合U2 snRNA，參與pre-mRNA splicing機制。表現夠量的OsRH42可保持pre-mRNA的splicing有足夠效能，並幫助水稻適應冷逆境，但OsRH42數量過多或過少，都會讓水稻植株中pre-mRNA splicing 不正常。因此，水稻植物需要精確控制OsRH42的表現量以反應環境溫度變化。雖然我們已了解OsRH42參與水稻對抗冷逆境之部分機制，但研究OsRH42影響之pre-RNA mRNA splicing是否具專一性及OsRH42是否需共同合作因子來參與調控冷逆境誘發pre-mRNA splicing的改變，對於進一步了解水稻受冷影響pre-mRNA splicing 之分子機制甚為重要。同時，以誘導啟動OsRH42來培育抵抗冷逆境的水稻，也是本研究計畫重點之一。本計畫因而具兩項主軸: 1. 基礎研究-水稻受冷逆境反應之分子機制的探討及2. 農業生物科技的開發-培育抵抗冷逆境的水稻。 ;Rice (Oryza sativa) is among the most important crops in the world, and is the main staple food for almost 50% of the world’s population. As a tropical and subtropical crop plant, rice is sensitive to cold stress. RNA molecules, including mRNA, rRNA, and tRNA, perform a specific function based on their well-defined structure during gene expression. Low temperature causes over-stabilization of incorrectly folded RNA and therefore leads to RNA molecular inactivation. RNA chaperones and RNA helicases function to ensure formation of mature RNAs of the correct structure by means of their RNA unwinding and RNA unfolding activities. RNA helicases are enzymes that can rearrange ribonucleoprotein (RNP) complexes and modify RNA structures, and are therefore involved in all aspects of RNA metabolism. DEAD-box helicases, which constitute the largest family of RNA helicases, exhibit variable protein sizes and compositions of N- and C-terminal extension sequences. Previously, we demonstrate that the rate of pre-mRNA splicing is reduced in rice at low temperatures, and the DEAD-box RNA helicase 42 (OsRH42) is necessary to support effective splicing of pre-mRNA during mRNA maturation at low temperatures. OsRH42 expression is tightly coupled to temperature fluctuation, and OsRH42 is localized in the splicing speckles and interacts directly with U2 snRNA. Retarded pre-mRNA splicing and plant growth defects were exhibited by OsRH42-knockdown transgenic lines at low temperatures, thus indicating that OsRH42 performs an essential role in ensuring accurate pre-mRNA splicing and normal plant growth under low ambient temperature. Unexpectedly, our results show that OsRH42 overexpression significantly disrupts the pre-mRNA splicing pathway, causing retarded plant growth and reducing plant cold tolerance. Combined, these results indicate that accurate control of OsRH42 homeostasis is essential for rice plants to respond to changes in ambient temperature. In addition, our study presents the molecular mechanism of DEAD-box RNA helicase function in pre-mRNA splicing, which is required for adaptation to cold stress in rice. However, it is important to understand the detail molecular mechanism of the OsRH42 function in cold stress resistance in rice. Therefore, the interaction proteins of the OsRH42 and the pre-mRNA targets of OsRH42 will be elucidated. Furthermore, inducible overexpression of OsRH42 in transgenic rice plants will be tested for their adapt ability under cold stress. Combining the results from these studies, we will have knowledge in OsRH42-mediated cold tolerance pathway and to develop a cold stress tolerant rice plants. |
