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標題: 水稻根部於鹽逆境下泛素修飾蛋白質體分析
Proteomic analysis of salt-responsive ubiquitin-related proteins in rice root
作者: 鄭宇翔
Cheng, Yu-Hsiang
關鍵字: rice;水稻;ubiquitin;salt tolerance;泛素;耐鹽
出版社: 分子生物學研究所
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目前全世界約有高於三分之一的人口以水稻作為主食,而水稻的生長與產量會受到鹽逆境的限制。由於全球土壤鹽化之問題日趨嚴重,因此如能藉由了解水稻於鹽逆境下之耐鹽反應,增進水稻之耐鹽特性,甚至增加水稻於鹽化土地上之產量將會是一個重要的課題。蛋白質的泛素 / 類泛素修飾在調節生物生理功能上扮演重要的角色,研究發現蛋白質不同之泛素 / 類泛素修飾位置、修飾型態與修飾數量,參與不同的生物生理機制。近年來有越來越多的證據顯示泛素 / 類泛素修飾蛋白在高等植物之生理機制中扮演關鍵角色,例如木質部發生、老化、細胞週期調控與逆境反應等。此外研究發現,在阿拉伯芥的RING finger E3 ligase (SALT- AND DROUGHT-INDUCED RING FINGER1, SDIR1) 參與離層酸之逆境訊息傳遞途徑,大量表現SDIR1會使離層酸所誘導之基因表現提高,例如提升萌芽時鹽的感受度、加強離層酸誘導氣孔關閉與增加乾旱的耐受性等。因此有效的鑑定出泛素 / 類泛素修飾之情況,將有助於研究泛素 / 類泛素修飾於植物生理調節所扮演的角色。本研究利用蛋白質體學與質譜技術,建立水稻泛素修飾蛋白質之分析平台。為研究泛素修飾蛋白質在鹽逆境反應中所扮演之角色,本實驗將栽培19天之水稻幼苗以250 mM的氯化鈉進行鹽逆境處理0.5、2、3與6小時。之後萃取根部蛋白質,再分別以膠體電泳與西方墨點轉漬法進行分析。從西方墨點轉漬法的結果發現,不同耐鹽特性之水稻根部其泛素修飾蛋白質表現有差異。本實驗以液相層析串聯質譜儀進行泛素修飾差異蛋白質分析,鑑定出蛋白質之身分,與泛素修飾蛋白質中泛素修飾之位置。從實驗結果推測這些泛素修飾蛋白質可能與水稻耐鹽特性有關,希望此研究成果能幫助了解泛素修飾參與水稻鹽逆境訊息傳遞相關蛋白之分子機制,提供日後耐鹽品種水稻育種之參考,進而提高鹽化土地水稻之栽培與產量。

Rice serves as a staple food source for more than one-third of the world's population. Rice growth and productivity is limited by salt stress. Because the global extent of soil salinity is increasing, it is important to understand the stress response of rice salt resistance for improving rice salt tolerance, especially for improving agricultural productivity in irrigated land. The ubiquitination / sumoylation of proteins play an important role in regulate function of physiology. Different types of ubiquitin (Ub) / SUMO conjugates are involved in the regulation of different cellular processes. In plants, ubiquitin is involved in a number of processes such as xylogenesis, senescence, cell cycle control and stress responses. In Arabidopsis thaliana, the RING finger E3 ligase, SALT- AND DROUGHT-INDUCED RING FINGER1 (SDIR1), is involved in abscisic acid (ABA)-related stress signal transduction. Over expression of SDIR1 leads to enhance ABA-induced stomatal closing, and enhanced drought tolerance. Therefore, efficient analysis of ubiquitin / ubiquitin-like proteins (Ubls) modifications will provide important information for their specific roles of cellular functions. In this work, we aim to develop analytical platform for Ub modifications in rice proteins analysis by using mass spectrometry (MS) combined with proteomic approaches. To investigate the role of ubiquitinaed proteins in the salt stress response, 19-days-old rice (Oryza sativa) seedlings were treated with 250 mM NaCl for 0.5, 2, 3, or 6 h. Total proteins of roots were extracted for comparative analysis compared with untreated controls using SDS-PAGE and western blot. In western blot, the results show that amount of ubiquitin-conjugated proteins were different between salt-tolerant and salt-sensitive rice seedling roots. These proteins were analyzed by LC-MS/MS, and the ubiquitination sites were also identified. These ubiquitin-conjugated proteins may be helpful in elucidating the molecular basis of salt tolerance in plant.
其他識別: U0005-1708200917552200
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