阿拉伯芥atToc33與atToc34基因之功能性研究

Functional studies of translocon genes atToc33 & atToc34 in Arabidopsis thaliana

陳怡彰(Yi-Jhang Chen)

指導教授 ： 孫智雯

國立臺灣師範大學/理學院/生命科學研究所/碩士(2009年)

摘要

葉綠體是植物行光合作用的重要胞器，在遠古時代由藍綠菌與真核細胞內共生而來。然而在演化過程中，超過90%的葉綠體蛋白質基因已經轉移至真核細胞的細胞核內。這些基因產物必須在細胞核內轉錄、在細胞質內轉譯及修飾後再送到葉綠體內。位於葉綠體膜上的轉運蛋白機組(translocon complex)負責辨識及運輸這些蛋白質，其中包含位在外膜上的Toc和位在內膜上的Tic兩大複合體。在阿拉伯芥中已知的轉運機組成員裡，atToc33和atToc34是豌豆Toc34的兩個同源蛋白，在運輸蛋白質進入葉綠體的過程中，扮演最初辨識蛋白質的角色。轉運機組的成員其基因必須被適當的調節，否則這些轉運機組將無法準確的組裝以執行適當的功能。為了了解atToc33和atToc34基因表現的調節的機制，atToc33和atToc34基因的啟動子序列接上GUS報導基因後送入野生型阿拉伯芥，結果顯示這兩個基因5’端非轉譯區內第一內插子對於促進基因表現有顯著的影響。此外，5’端啟動子刪減及膠體電泳位移分析實驗亦找出數個可能調節atToc33基因表現的啟動子區域。

關鍵字

atToc33 ； atToc34

並列摘要

Chloroplast, originated from endosymbiotic cyanobacteria in the ancient times, is one of the most important organelles in plants for it’s function of photosynthesis. During the evolution of plant cells, more than ninety percent of the plastid genes were gradually transferred to the host nuclear genome. However, these genes have to be transcribed in nucleus, translated and post-translationally modified in cytoplasm, and imported to plastids accurately. The translocons on the outer envelope and inner envelope membrane of chloroplast play key roles on machinery of protein import into the chloroplast. They compose two complexes: Toc complex (Translocon at the Outer envelope membrane of the Chloroplast) and Tic complex (Translocon at the Inner envelope membrane of the Chloroplast). Among the Toc and Tic components identified in Arabidopsis thaliana, atToc33 and atToc34 are homologous to pea psToc34 and play important roles of pre-protein recognition. Evidently, the expression of translocon genes have to be regulated properly, or their gene products will not be accurately integrated to their destination and proceed their mission. In order to reveal the regulatory mechanism of atToc33 and atToc34 gene expression, transgenes containing the promoter sequences of atToc33 and atToc34 gene and GUS coding sequence were transferred into wild-type Arabidopsis. The 1st intron in the 5’UTR of both gene have obvious effect on the regulation of gene expression. Furthermore, 5’ promoter deletion and EMSA experiments reveal several putative regulation regions of promoter sequence to modulate atToc33 expression.

並列關鍵字

atToc33 ； atToc34

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