香蕉ABC轉運蛋白基因之生理及生化功能分析

Abstract

為了瞭解香蕉ABC轉運蛋白MhPDR1及MhPDR2在生理及生化的功能，本研究針對過量表達香蕉ABC轉運蛋白的轉殖株Nt-MhPDR1及Nt-MhPDR2進行試驗。以不同濃度之重金屬銅、鎘和鋅處理，再進行生理生化及植體內重金屬含量分析，結果顯示，與未轉殖株相比，轉殖株對銅、鎘和鋅離子有較高的抗性，進一步分析顯示，轉殖株體內含較低量的重金屬銅、鎘和鋅離子，且體內總葉綠素含量較未轉殖株高，轉殖株受脂質過氧化程度較未轉殖株低，因此推測MhPDR1及MhPDR2扮演對重金屬銅、鎘和鋅離子輸出幫浦 (efflux pump) 的角色。另一試驗以營養元素氮素、磷酸、鉀和鐵離子進行處理，結果顯示，轉殖株體內營養元素含量都高於未轉植株，因此推測MhPDR1及MhPDR2扮演對大量元素運移的角色。而為了瞭解ABC轉運蛋白MhPDR1及MhPDR2在細胞中表現的位置，將MhPDR1及MhPDR2分別與綠色螢光蛋白 (green fluorescent protein, GFP) 基因融合，構築為表現載體，利用PEG法轉殖到阿拉伯芥 (Arabidopsis thaliana L. ecotype Columbia) 之原生質體 (protoplast) ，透過觀察融合蛋白 (fusion protein) 的位置，了解香蕉ABC轉運蛋白在植物中表達位置，結果顯示MhPDR1位在液胞膜，而MhPDR2則是位在細胞膜上。

In order to understand the function of banana ABC transporter MhPDR1 and MhPDR2, ABC transporter (MhPDR1 or MhPDR2) overexpressed tobacco plants (Nt-MhPDR1 or Nt-MhPDR2) have been tested by several treatments. The results of heavy metals treatments indicated that Nt-MhPDR1 and Nt-MhPDR2 have higher tolerance to copper, cadmium, and zinc ions than untransformed plants. In heavy metal content analysis, the results showed that Nt-MhPDR1 and Nt-MhPDR2 accumulated lower concentrations of copper, cadmium, and zinc. In chemical component analysis, the results showed that Nt-MhPDR1 and Nt-MhPDR2 have higher chlorophyll and lower lipid peroxidation than untransformed plants. Therefore, banana ABC transporter MhPDR1 and MhPDR2 are suggested to play role as efflux pump of the copper, cadmium, and zinc ions. Furthermore, transgenic tobacco Nt-MhPDR1 and Nt-MhPDR2 accumulated higher nitrate, phosphate, potassium, and iron contents than untransformed plants. Therefore, MhPDR1 and MhPDR2 may also possibly play the role as a transporter for these elements. To investigate the cellular localization of ABC transporter MhPDR1 and MhPDR2, coding regions of MhPDR1 and MhPDR2 were fused with the green fluorescent protein (GFP) gene and transiently introduced into Arabidopsis protoplasts by PEG method. The result indicated that MhPDR1 and MhPDR2 are located at tonoplast and plasma membrane, respectively.