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)的長度同時影響mRNA穩定度與蛋白質轉譯作用，因此調控mRNA tail長度是影響基因表現的重要環節。mRNA poly(A) tail deadenylation是許多mRNA降解的速率決定步驟，負責真核細胞內deadenylation作用機制的主要蛋白質為CCR4-CAF1複合體。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，但對於植物與水稻中的CCR4與CAF1所知甚少。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。其中OsCAF1B在水稻細胞內確實扮演deadenylase的角色，它參與調控水稻糖訊息誘導αAmy3 mRNA deadenylation以及αAmy3 mRNA穩定度。同時我們也發現OsCAF1B為受低溫誘導表現的基因並參與低溫逆境相關路徑，具有deadenylase 活性的OsCAF1B大量表現後，可幫助水稻轉殖株度過冷逆境。然而，OsCAF1B卻無法與OsCCR4a及OsCCR4b結合，所以OsCAF1B可能自己執行deadenylation功能，雖然OsCAF1B與NOT也會結合，形成complex的成員結合也可能有所差異。因此，延續先前研究計畫，以現有水稻轉殖株為基礎，完整與確切釐清OsCAF1B參與水稻對抗冷逆境的分子機制。同時，由於我們已發現大量表達OsCAF1B的水稻植株具有耐冷逆境的良好應用性，但大量表達OsCAF1B卻影響水稻產量，因此利用誘導性啟動子來培育耐冷逆境的水稻也是本研究計畫重點之一。 ;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 two components CCR4 and CAF1, is a major player in deadenylation. Moreover, the CAF1 have been hypothesized that plays important roles to cope with various environmental stresses in plants. Previously, we identified 4 CAF1 genes, OsCAF1A, OsCAF1B, OsCAF1G and OsCAF1H, from rice that encode recombinant proteins all exhibited 3’ to 5’ exonuclease activity in vitro and tended to remove poly(A) tails. One of them, OsCAF1B, have demonstrated that plays a vital role in sugar-induced αAmy3 mRNA degradation and deadenylation. Our study presents also show that expression of the OsCAF1B gene was cold induced and overexpress the active form of OsCAF1B enhanced cold tolerance phenotype in rice. These results imply that the OsCAF1B play a critical role for deadenylation of cold-responsive gene mRNAs and contribute to rice tolerance to cold. This proposed research focus to study the detail molecular mechanism of the OsCAF1B in cold stress responses in rice. Therefore, the interaction proteins of the OsCAF1B and direct cold-stress related target mRNAs of OsCAF1B will be elucidated. Meanwhile, inducible overexpression of OsCAF1B in transgenic rice plants will be tested for their adaptability under cold stress. Combining the results from these studies, we will have enough knowledge in OsCAF1B-mediated cold tolerance pathway to develop cold stress tolerant rice plants.
