T-DNA的嵌入干擾At3g22970基因表現且使阿拉伯芥的葉部形態異常

葉，植物感受環境變化的營養器官，對光合作用、呼吸作用和氣體交換等生理作用有著重要角色，然而到目前為止對於葉部發育細節仍不是十分清楚。實驗室利用T-DNA載體，pPZP202: SK: BAR，隨機嵌入阿拉伯芥基因體中的方法建構了突變株庫，篩選葉部形態異常的突變株並尋找影響此性狀的基因。R1-2突變株，為本實驗室所篩選一株具有捲曲葉形性狀的突變株，已知T-DNA嵌入在At3g22970上游約2.5-Kbp處且At3g22970的基因表現量下降。本研究，則對At3g22970在植物生理上的功能及T-DNA如何影響基因表現做進一步的研究。實驗結果顯示，At3g22970的基因表現在wild type (WT)中具有日夜週期性(circadian rhythm)，於近夜間時期(early-night phase)有最大表現量。但在R1-2植株中，At3g22970的基因表現量大幅下降但仍觀察到有日夜週期性的表現，因此推測T-DNA嵌入在At3g22970上游約2.5-Kbp處可能影響到增強子(enhancer)對基因表現的正向調控。At3g22970是個功能未知的葉綠體蛋白，結構上具有一個高保留性的DUF506區域 (Domain of Unknown Function 506) 並可送入質體。在阿拉伯芥中，共有14個基因含有此區域且功能都尚未被研究。我們利用At3g22970的啟動子表現報導基因，結果顯示在較年輕的組織中有較高的表現量。且在不同的過量表現At3g22970的植株中，葉部形態也展現不一致性狀，推論其表現量對植物發育可能有劑量性的影響。另外，我們利用全長的At3g22970以Yeast Two-Hybrid方法篩選出9個與At3g22970有交互作用的蛋白，已知其分別參與於光合作用，醣類代謝，訊息傳遞以及轉譯作用等。這些與At3g22970有交互作用的蛋白，我們將做更進一步的實驗去探討它們與At3g22970的關係，以幫助我們了解At3g22970蛋白在植物生理上的功能。

關鍵字

捲曲葉形

並列摘要

The frontier of leaf morphogenesis is still undiscovered, particularly in the dicotyledonous plants. Therefore, using a T-DNA tagging vector, pPZP202: SK: BAR, to generate mutant pools. Searching for plants with abnormal leaves phenotype and identify the genes which influenced by T-DNA in Arabidopsis thaliana. In previous study, we have identified a curly leaf mutant, ‘R1-2’. T-DNA inserts in 2.5-Kbp upstream of the At3g22970 and At3g22970 gene expression was down-regulated in the R1-2. In this study, we focused on how and why T-DNA insertion interfered with At3g22970 gene expression in R1-2 to result the phenotype. At3g22970 showed a circadian rhythm expression and the most expression during the subjective dusk. The transcripts level of At3g22970 in R1-2 presented all down-expression and had a slight circadian rhythm expression that was compared to WT. We propose that T-DNA insertion may disrupt the function of some enhancer for promoting At3g22970 gene expression in R1-2. At3g22970 encodes a protein containing a highly conserved DUF506 domain (Domain of Unknown Function 506) and translocated into plastid; however function still unknown. Meanwhile, there are 14 genes which containing DUF506 domain and all shown unknown function in A. thaliana. Moreover, At3g22970 promoter-GUS assay showed that At3g22970 mainly expressed in younger tissues. Overexpression of At3g22970 shown different degree interfere with leaf morphology suggested that At3g22970 function with dosage-effect. Furthermore, yeast two-hybrid results showed that At3g22970 interacts with 9 different proteins which involved in photosynthesis, carbohydrate metabolism, signal transduction and translational regulation. The relationships between At3g22970 and the interacting proteins need further study to understand the physiological function of At3g22970 in the future.

並列關鍵字

curly leaf ； circadian rhythm ； DUF506

參考文獻

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T-DNA的嵌入干擾At3g22970基因表現且使阿拉伯芥的葉部形態異常

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