| 摘要: | 高等陸生植物(包含水稻)時常必須面對各種環境逆境壓力，為了提高生存能力就必須適時改變自身 的分子運作及生理機轉，包含誘導或抑制相關基因的表現，以便能渡過逆境所造成的生存壓力。mRNA poly(A) tail deadenylation 是許多mRNA 降解的速率決定步驟，藉由poly(A) tail deadenylation 引起 mRNA 的降解使mRNA 含量降低，mRNA poly(A)的長度同時也影響蛋白質轉譯作用，因此調控mRNA tail 長度是影響基因表現的重要環節。CCR4-CAF1 複合體為真核細胞內負責deadenylation 作用機制的 主要蛋白質複合體且具有高度保留性，然而這些研究大多著重在動物或酵母菌，對於植物與水稻中的 CCR4-CAF1 則所知甚少。 CAF1 與CCR4 是CCR4-CAF1 複合體中負責poly(A) tail deadenylation 的主要次單元，CAF1 在高 等真核生物中尤其重要。過去少數植物CAF1 的文獻報告指出，植物的CAF1 具有deadenylase 的活性， 而且影響植物對環境逆境的耐受能力，唯該方面的研究，主要集中在雙子葉植物，如：阿拉伯芥與辣 椒等植物。先前我們發現水稻有OsCAF1A、OsCAF1B、OsCAF1G 與OsCAF1H 4 個CAF1 基因，它們 均擁有deadenylase 活性可執行deadenylation 功能，並各有獨特的生理功能，其中OsCAF1B 與OsCAF1H 可能分別參低溫與高溫逆境相關路徑，並可能調控了部分基因的表現。因此，本延續研究計畫的主要 目的以現有水稻轉殖株為基礎，釐清(1) OsCAF1B 參與水稻抗冷逆境的調控機制。(2) OsCAF1H 參與 水稻抗熱逆境的調控機制。 本計畫將分別釐清OsCAF1B 與OsCAF1H 在冷、熱環境逆境中如何調控下游基因mRNA poly(A) 長度的分子機制，以及mRNA deadeylation 在mRNA 穩定度及mRNA 累積量的重要性，一窺水稻如 何藉由不同的調控階段，在適當的在水稻對抗冷和熱逆境時適量的表現特定的基因，這些知識有助於 全盤瞭解水稻生長發育與環境逆境反應在後轉錄調控基因表現之分子機制，將來可應用於提高水稻環 境抗逆境之農業生物科技上。 ;Plants are sessile require complex and coordinated gene expression to survive under stress environments. Deadenylation, also called poly(A) tail shortening, disrupts the circularized mRNP structure, which in turn suppresses translation and commits the mRNA to degradation. Thus, deadenylation is an important step in gene regulation that controls mRNA stability and translation efficiency. The CCR4-NOT complex, containing the 2 key components CCR4 and CAF1, is a major player in deadenylation. It is plausible to hypothesize that the CAF1 play important roles in plants to cope with adverse conditions in Arabidopsis. However, little is known whether crop plants employ similar mechanisms to overcome stressful environments and whether manipulation of CAF1 deadenylase activities could enhance crop tolerance. This proposed research focus to study the roles of two CAF1, OsCAF1B and OsCAF1H, for environmental stress resistance in rice (Oryza sativa). Previously, we have identified 4 CAF1 genes, OsCAF1A, OsCAF1B, OsCAF1G and OsCAF1H, which all exhibited 3’ to 5’ exonuclease activity in vitro and tended to remove poly(A) tails. The unique expression and subcellular localization patterns of each OsCAF1 were observed in various tissues when undergoing abiotic stress treatments, implying that each CAF1 gene plays a specific role in the development and stress response of a rice. Expression of the OsCAF1B gene was cold induced and the OsCAF1B overexpression transgenic lines exhibited cold tolerance phenotype. On the other hand, OsCAF1H is a heat upregulated gene, OsCAF1H RNAi line showed a heat insensitive phenotype, whereas OsCAF1H overexpression transgenic seedling displayed tolerance to heat stress. Moreover, the expression of small heat shock protein gene, OsHsp18.0, showed the different pattern in OsCAF1H RNAi knockdown line compared to wild type. These results imply that the OsCAF1B and OsCAF1H, play a critical role for deadenylation of cold- and heat-responsive gene mRNAs and contribute to rice tolerance to cold and heat stresses, respectively. This proposed research focus to study the detail molecular mechanism of the OsCAF1B and OsCAF1H functions in temperature stress responses in rice. Therefore, the functions of the OsCAF1B and OsCAF1H in physiological, cellular and molecular responses to a variety of temperature stresses and the targets of OsCAF1B and OsCAF1H will be elucidated. Furthermore, overexpression of OsCAF1B and OsCAF1H in transgenic rice plants will be tested for their adaptability under temperature stress. Combining the results from these studies, we will have enough knowledge in OsCAF1B and OsCAF1H to develop cold and heat stresses tolerant rice plants, respectively. |
