小花豬籠草銨轉運蛋白功能分析

Functional Characterization of Nepenthes graciliflora Ammonium Transporter, NgAMT1

江恆安(Heng-An Chiang)

指導教授 ： 王雅筠

國立臺灣大學/生命科學院/生命科學系/碩士(2022年)

https://doi.org/10.6342/NTU202203736

摘要

氮是植物生長所不可或缺的元素，植物透過硝酸鹽或銨鹽做為主要無機氮源，而銨轉運蛋白是植物吸收銨鹽重要的途徑。豬籠草是一類分布於缺氮環境的食肉植物，透過將獵物身上的蛋白質分解為銨鹽等小分子以吸收動物體內的養分，透過研究其銨轉運蛋白能了解於缺氮環境下植物所演化出的適應機制與非食肉植物有何差異，以期對提升植物氮利用效率有所幫助。過去研究中發現假定為翼狀豬籠草銨轉運蛋白NaAMT1的mRNA專一性表現於捕蟲瓶內腺體細胞，並會受到獵物落入的誘導增加，本研究透過RACE PCR自小花豬籠草(Nepenthes graciliflora, 原翼狀豬籠草Nepenthes alata同物異名) 捕蟲瓶RNA中取得NgAMT1 CDS全長。在酵母菌表現系統中，NgAMT1於pH 4.0下未表現出銨離子轉運功能；在蛙卵母細胞表現系統中，於pH 5.5環境中亦無觀察到吸收銨離子功能。在酵母及菸草表現系統中，NgAMT1於細胞中之表現位置均在內質網。另觀察到表現NgAMT1之酵母於pH 6.6環境下，生長較空載體控制組顯著遲緩。推測該轉運蛋白可能參與內膜系統之銨離子運輸，其詳細功能尚待更多研究。

關鍵字

食肉植物 ； 豬籠草 ； 銨轉運蛋白 ； AMT ； NaAMT1 ； NgAMT1

並列摘要

Nitrogen is an essential element for plant growth. Plants absorb inorganic nitrogen as ammonium or nitrate. Ammonium transporters (AMTs) contribute most of the ammonium influx from the soil. Nepenthes is a genus of carnivorous plants distributed in nitrogen-poor environments. By digesting the protein of the prey into small molecules like ammonium, Nepenthes utilize the nutrients from the animals. Studying how they uptake ammonium may understand how plants adapted to nitrogen-poor environments and thus sheds light on improving nitrogen use efficiency (NUE). In previous research, a putative NaAMT1 mRNA is detected explicitly in Nepenthes alata pitcher gland cells. The expression of NaAMT1 is upregulated by the prey. In this study, the full-length CDS of NgAMT1 was cloned from the total RNA of the pitcher of Nepenthes graciliflora (synonym of Nepenthes alata until 2013) by RACE (Rapid amplification of cDNA ends) PCR. NgAMT1 exhibits no ammonium uptake activity in the yeast-expressing system at pH 4.0. In the oocytes-expressing system, NgAMT1 shows no ammonium uptake ability at pH 5.5. In the subcellular localization, the NgAMT1 was observed in the endoplasmic reticulum (ER) in yeast cells and tobacco leaf epidermis cells. Surprisingly, the NgAMT1-expressing yeasts showed growth retardation at pH 6.6 compared to empty vector control, suggesting that NgAMT1 probably participates in ammonium transportation in endomembrane systems. Further research is required to examine the function of NgAMT1 in pitcher plants.

並列關鍵字

carnivorous plants ； Nepenthes ； AMT ； NaAMT1 ； NgAMT1

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