Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/31747
標題: 生物及非生物逆境對第一族低分子量熱休克基因疾病程相關基因的誘導
Induction of class I low moleculer mass heat shock protein genes and a pathogenesis-related gene in rice by biotic and abiotic stresses
作者: 張喜民
Hsi-Sming, Chang
關鍵字: 逆境
熱休克基因
病程相關基因
水稻
出版社: 植物病理學系
摘要: 熱休克蛋白質 (heat shock proteins; HSPs) 存在於很多生物體中,目前已知環境逆境 (stresses) 會誘導這些蛋白質的產生,這些蛋白質的功能大多與受到環境逆境時用來維持其它蛋白質結構的穩定性或是分解一些變性蛋白質有關, HSPs 在植物也有類似的保護效果,例如:在植物中含量特別多的低分子量熱休克蛋白質 (low molecular mass HSPs, LMM HSPs) ,其功能也是提供植物在逆境下的耐性。另一方面,病程相關 (pathogenesis-related, PR) 基因和「系統性後天抗性」 (systemic acquired resistance, SAR) 在植物對病原菌的防禦機制上扮演一個很重要的角色,有多篇報告指出若在轉基因植物體內大量表現一些 PR 基因會使植物對病原菌的攻擊有耐性。本研究利用高溫 41℃ 處理、水楊酸 (salicylic acid) 及切傷處理 3 天大水稻未照光幼苗,藉以偵測不同處理對水稻第一族 (class I) LMM HSP 基因及 PR-1 基因的誘導。結果顯示,第一族 LMM HSP 基因不僅可被熱休克誘導也可被水楊酸及切傷處理所誘導,但其基因表現程度較熱休克處理低。另外,目前已知在水稻中至少有 6 個以上之第一族 LMM HSP 基因,利用專一性「聚合酵素鏈反應」(polymerase chain reaction, PCR) 的引子對可分別偵測單一基因之表現,由 「反轉錄-聚合酵素鏈反應」(reverse transcription-PCR, RT-PCR) 之結果顯示各處理對誘導不同第一族 LMM HSP 基因之表現量也有差異,而且在基因表現量方面,切傷處理 4 小時對於 Oshsp18.0 的誘導量比對照組高 1.62 倍,與 41℃ 熱休克所誘導的 1.8 倍相近。另各處理樣品之總蛋白質以第一族 LMM HSP 之多株抗體進行西方墨漬分析 (western blot analysis) 其結果同樣也顯示熱休克處理可大量累積 class I LMM HSP,而水楊酸處理也可誘導其蛋白質累積,只是程度較低。另以 RT-PCR 以及北方雜合法 (northern hybridization) 分析之結果顯示:在上述處理下,水稻病程相關基因 PR-1 的表現並沒有太大的差異。由於熱休克蛋白質除了提高生物的耐熱性並保護細胞內蛋白質在逆境下不致變性外,在動物系統中其功能也和組織發炎有關,而且在胞囊線蟲 (Heterodera glycines Ichinohe) 感染大豆時,HSP70 基因的表現量也會提高,推測其在植物-病原菌交互作用中也可能扮演相關角色,故進一步利用立枯絲核菌 (Rhizoctonia solani R96) 接種 1 個月大的水稻植株以分析其第一族 LMM HSP 及 PR-1 基因在這些生物逆境下之表現。結果顯示病程的發展和 PR-1 基因的表現有很大的關係,此外,以 R. solani R96 接種水稻植株可分別誘導第一族 LMM HSP 基因中的 Oshsp16.9B,Oshsp16.9C,Oshsp17.0推測在整個植物對抗病原菌侵入的過程中,HSP 基因的表現也應和 PR 基因一樣扮演某種相關的角色。
HSPs are produced in microorganisms, plants and animals to protect tissues from high temperature stress. The functions of HSPs were reported in many living organisms. Low molecular mass HSPs (LMM HSPs, or small HSPs, smHSPs) are especially abundant in plants and play a major role in thermoprotection. On the other hand, pathogenesis-related (PR) proteins are associated with systemic acquired resistance (SAR) in plant defense responses. Overexpressing the PR genes may protect transgenic plants from pathogen attacks. A number of genes, including THE HSP70 family, were induced in soybean upon infection with cyst nematode. In animal system, HSPs are also related with inflammation but no report has been presented to connect HSPs with pathogen resistance in plants. The objective of this study was to investigate the expression of class I LMM HSP genes and PR-1 gene under biotic and abiotic stresses in rice seedlings and plants. The class I LMM HSP genes and PR-1 gene expression in rice seedlings treated with 41C, salicylic acid, or wounding, and in rice plants inoculated with Rhizoctonia solani R96 was observed in this study. Among the six rice class I LMM HSP genes tested, the Oshsp18.0 expression was induced by 4 h of wounding treatment to a level of about 1.62-fold higher than that by control treatment, similar to that of 1.8-fold by 41C HS treatment, according to the results of reverse transcription (RT)-polymerase chain reaction (PCR). The results of western blot analysis using polyclonal antibodies specific to the class I LMM HSPs also suggested that 4 h of salicylic acid treatment induced the accumulation of class I LMM HSPs. However, no much expression of the rice PR-1 gene was induced by the above-mentioned abiotic treatments as suggested by the results of RT-PCR and northern hybridization. On the other hand, PR-1 gene expression was induced, and related to pathogenesis by Rhizoctonia solani R96 infection. Four class I LMM HSP genes Oshsp16.9B, Oshsp16.9C, and Oshsp17.0 were induced by Rhizoctonia solani R96 infection. The results here suggested that, beside the PR proteins, the HSPs may also play a role in plant defense mechanism against the pathogen attack.
URI: http://hdl.handle.net/11455/31747
Appears in Collections:植物病理學系

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