噬菌體裂解蛋白與青枯病菌gpsA對番茄青枯病反應之影響

Effects of a phage lysis protein and Ralstonia solanacearum gpsA on tomato bacterial wilt response

黃琪(Chi Huang)

指導教授 ： 鄭秋萍

國立臺灣大學/生命科學院/植物科學研究所/碩士(2015年)

https://doi.org/10.6342/NTU.2015.10081

摘要

植物經常會受到各種不同病原菌攻擊，導致嚴重作物生產損失，例如：由青枯病菌所引起的青枯病，即是全世界最嚴重的植物病害之一，影響全球許多重要經濟作物，造成極大損失。本研究包含兩個子題。在第一子題中，我們先前的研究顯示，在菸草中表現源自青枯病菌之噬菌體的一個裂解蛋白質H會造成被細菌感染誘導的植物細胞死亡，並使植物對數種病原細菌之抗性增加。本研究進一步證實，在番茄植株系統性表現H蛋白質可加強植物抵抗多種特性不同之細菌病害，亦會造成細菌誘發性細胞死亡與增強pathogen-associated molecular patterns (PAMPs)-triggered immunity (PTI) 反應，故表現H蛋白質可在植物體內誘導廣泛性防禦反應而促使植物得以抵抗不同的病原菌。在第二子題中，我們先前的研究顯示，青枯病菌參與糖解作用之酵素基因gpsA經破壞後，會導致病菌在番茄抗病品系中之毒力下降。本研究進一步證明gpsA確實是決定青枯病菌致病力的關鍵基因，並參與病菌之糖利用作用；同時，定量分析顯示番茄感病番茄品系L390之葡萄糖、果糖及蔗糖含量較抗病品系H7996高，且在L390根部能測到低量甘油 (glycerol) 而在H7996之各部位中均無法測得甘油；此外，L390在感染gpsA突變株後，其根部中之甘油含量下降趨勢比感染野生型菌株快速。這些結果顯示，在甘油含量低至無測得之抗病番茄品系H7996中，gpsA在青枯病菌感染的初期扮演重要角色，使病菌得以在甘油含量極低之H7996根部增殖，以達成後續的系統性感染，同時，這些發現也暗指番茄之含糖量可能與其青枯病抗性具一定相關性。另外，從本研究之番茄與馬鈴薯糖類傳輸蛋白 (sugar transporters) SWEETs序列分析結果發現，茄科植物之SWEETs基因可能比阿拉伯芥與水稻略多，且短暫基因靜默分析初步發現番茄SWEET2a (SISWEET2a) 參與植株生長發育。預期本研究所獲資訊將有助於瞭解植物病害發生之具體機制，並研擬防治病害之可能措施。

關鍵字

番茄 ； 青枯病菌 ； 噬菌體 ； 裂解蛋白質 ； gpsA ； 糖類 ； SWEET

並列摘要

Plants often suffer from attacks of different pathogens, causing serious losses in crop production. For example, bacterial wilt (BW), caused by Ralstonia solanacearum (Rs), is a serious disease affecting a wide range of economically important crops worldwide. This work contains two studies. Firstly, our previous studies revealed that expression of a lysis protein (H) derived from a bacteriophage of Rs promoted bacteria-induced cell death and resistance to several bacteria in Nicotiana benthamiana. Results from this study further revealed that systemic overexpression of H or signal peptide (sp)-fused H proteins enhanced tomato resistance against three distinct bacteria, bacteria-triggered cell death and pathogen-associated molecular patterns (PAMPs)-triggered immunity (PTI) responses. Therefore, expression of H protein can prime plants for enhanced defense responses, leading to boosted resistance to different pathogens. Secondly, our previous studies revealed that the disruption of Rs gpsA, which encodes an enzyme involved in glycolysis, led to differential bacterial virulence in BW-resistant and -susceptible tomato cultivars. Data from this study further evidence that gpsA is a general virulence determinant of Rs and is involved in sugar utilization. In addition, contents of sugars (glucose, fructose, sucrose) in BW-susceptible tomato cultivar L390 are higher than that in the resistant cultivar Hawaii7996 (H7996). The content of glycerol was detected at a very low level in the root of L390 but not in other tissues of L390, and glycerol was undetectable in various tissues of H7996. Moreover, the glycerol content in L390 root after the infection of Rs gpsA mutant decreased more dramatically than that after the infection of the wild-type strain. These results suggest that, to achieve a successful infection in H7996 with very limited glycerol, if there is any, functional gpsA is required for Rs sugary utilization and thus proliferation at the initial infection stage. In addition, these data imply an effect of sugar contents on tomato BW response. Furthermore, sequence analyses reveal a higher number of sugar transporters SWEET homologs in tomato and potato than that in Arabidopsis and rice. Moreover, transient gene silencing assay suggest a key role tomato SWEET2a (SISWEET2a) in plant growth. This work together with the proposed further studies are anticipated to provide important information on plant defense mechanisms and explore useful defense genes, potentially paving the way to assemble effective approaches for BW control.

並列關鍵字

tomato ； Ralstonia solanacearum ； bacteriophage ； lysis protein ； gpsA ； sugars ； SWEET

參考文獻

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