SAHH基因的分離及其對於番茄茄紅素累積之功能探討

Abstract

番茄是眾人在茶餘飯後常見的水果之一，除了作為水果外，也可以用來烹調成美味的佳餚，在研究上則屬於研究果實成熟的模式植物之一。它是一種需要經過後熟作用的更年性果實(climacteric fruit)，與香蕉一樣都是需要經過乙烯的催熟才能從綠熟(mature green)變成紅熟(red ripening；RR)的果實。乙烯在植物的生長發育過程中扮演極為重要的角色，從幼苗到成株再到花、果實種子發育等等，都有它的影響，甚至是逆境的調控也少不了它。在先前的研究指出，TDR4 (FUL1)這個MADS-box的轉錄因子(transcription factor)會與其他轉錄因子像是RIPENING INHIBITOR (RIN)形成複合體調控乙烯生合成路徑，進而調節番茄的紅熟過程；除了乙烯，也有研究指出亦能藉由非乙烯依賴(ethylene independent)的途徑，促使果實成熟，除了藉由其餘賀爾蒙外，光的影響也參與在其中。實驗室前人的研究中，發現藍光及紅光會使番茄的茄紅素累積的較快，且TDR4與S-adenosyl-L-homocysteine hydrolase (SAHH)這兩個基因表現也有顯著的提升，值得一提的是SAHH這個水解酶所參與的SAM循環恰巧也能夠調節乙烯上游的ACC生合成，顯然SAHH與TDR4之間有相當程度的調控關係。我的研究主要是建立並且探討SAHH2大量表現的轉殖株，並找出這兩者間在番茄茄紅素累積過程中的交互關係。本篇中以TDR4抗體偵測到植物體中的TDR4多半穩定表現在果實成熟階段中，其中又以Breaker stage所含最多，而在RR stage可能有其他的蛋白修飾存在。番茄當中有三個SAHH的同源基因，在果實各個階段的基因表現與TDR4呈現一個正相關，但其中又以SAHH2與TDR4關聯最大，另外SAHH2與JRL1之間的調控可能會藉由光的影響而有所不同。在番茄中大量表現SAHH2，會使得植株外型有所改變，造成較小且扭曲的葉片，也會使果實提早進入紅熟期並增加其對於乙烯的敏感性，由此可見，SAHH2不僅僅只參與在果實紅熟的調控中，對於植物的生長發育也有很大的影響。

Tomato is a famous fruit in the world. It’s not only a fruit, but also a vegetable. Many researchers use it as a model plant to study fruit ripening. Tomato is a climacteric fruit which turns the color of the exocarp from green to red due to ethylene effect. Ethylene participates in the regulation of plant development such as seedling, flowering, fruit. Many factors affect tomato fruit ripening, in which the process of ethylene plays a very important role. TDR4 is a transcription factor of the MADS-box family, which participates in the ripening process of tomato fruit, and can also form complexes with other transcription factors. This complex can regulate ethylene biosynthetic pathway, leading to the regulation of fruit ripening. Besides ethylene, many factors such as other hormones and light may affect fruit ripening. Previous studies showed that gene expressions of TDR4 and SAHH were up-regulated when lycopene accumulation was induced by blue light and red light. In addition to TDR4, the enzyme S-adenosyl-L-homocysteine hydrolase (SAHH) that associates with the SAM cycle may have a potential function to regulate tomato ripening. In my study, I wonder whether SAHH participates in the process of lycopene accumulation and may have a relationship with TDR4. Current results show that TDR4 protein is only expressed in tomato fruit and its levels peaked at the breaker stage. Moreover, there may be protein modification in red ripen stage. SAHH has three homologs in tomato. We examined their gene expressions and indicated that SAHH2 may play an important role in fruit ripening, especially in the breaker stage. Also, SAHH2 may have a regulatory relationship with JRL1 under different light conditions. In addition, the connection between SAHH2, JRL1 and TDR4 remains unclear. We have established SAHH2 overexpression transgenic tomato lines in order to study the regulatory relationship among these three genes. Current evidence reveals that SAHH2 overexpression transgenic lines exhibit curled shapes of leaflets and are sensitive to ethylene. Taken together, this study indicates that SAHH2 not only regulates fruit ripening, but also affects the plant growth and development.