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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 葉信宏 | |
| dc.contributor.author | Cheng-En Chen | en |
| dc.contributor.author | 陳正恩 | zh_TW |
| dc.date.accessioned | 2021-06-08T04:15:52Z | - |
| dc.date.copyright | 2010-08-10 | |
| dc.date.issued | 2010 | |
| dc.date.submitted | 2010-08-04 | |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/22342 | - |
| dc.description.abstract | 病毒入侵寄主之初期，和寄主所產生的交互作用常決定了病毒是否可在寄主中複製及侵染。為了瞭解病毒感染初期和寄主產生的交互作用，針對病毒感染初期所誘發的寄主轉錄體變化進行探討，以阿拉伯芥的cDNA microarray分析TMV複製早期寄主轉錄體的變化，發現到一寄主基因PAP85於一般發育階段無明顯表現，但在TMV感染初期其會被穩定的正向調控，而藉由PAP85的負向調控可對病毒的累積產生抑制效果，進一步研究顯示TMV可能會利用PAP85去影響寄主內質網的結構以促進病毒複製複合體的生合成。此外一轉錄因子WRKY6會於病毒感染後4小時被正向調控，先前之研究已知WRKY6參與了植物的抗性反應，而進一步於WRKY6大量表現或抑制表現的原生質體及植物中分析TMV的累積情況，結果顯示WRKY6雖然在病毒複製初期無顯著影響，但WRKY6於植物中同時扮演了促進和抑制TMV感染的角色。而為了瞭解組蛋白修飾過程是否亦參與在病毒感染初期和寄主產生之交互作用，我們針對組蛋白乙醯化酶(HATs)及組蛋白去乙醯化酶(HDACs)進行突變株的篩選及分析，結果發現到一CBP family的HAT基因(HAC2)於TMV在原生質體中的累積扮演重要角色。綜合這些結果顯示，在病毒感染初期和寄主產生交互作用時，寄主的基因表現、轉錄因子和組蛋白修飾過程皆可能會受到調控，而藉由控制這些被病毒調控的寄主因子也許可在病毒防治上提供可行的策略及方法。 | zh_TW |
| dc.description.abstract | The early stage of interaction between viruses and hosts is a detrimental step for successful virus replication and further infection. In order to understand the early stage of interaction during virus infection, the host transcriptome change in response to virus early infection was studied. Arabidopsis cDNA microarray was conducted to monitor host transcriptome change during the early stage of infection of Tobacco mosaic virus (TMV). A host gene, PAP85, with low expression level in most development stages but continuatively up-regulated in early stage of TMV infection was identified. Virus accumulation could be inhibited by down-regulation of PAP85. My data also indicated that TMV may adapt PAP85 to alter endoplasmic reticulum (ER) structure in facilitating the formation of virus replication complexes. A transcription factor (TF), WRKY6, which was up-regulated at 4 hour post virus infection was selected for further study. The WRKY6 TF was known to involve in plant resistance to pathogens in previous studies. TMV inoculation assay was conducted in WRKY6-overexpressed and -knockout protoplasts and plants. Although WRKY6 may not have the effect on the initial establishment of virus replication, WRKY6 plays both supportive and inhibitory roles in TMV accumulation. I also tried to understand whether histone modification are involved in early stage of virus infection. I performed mutant screening of histone acetyltransferase (HATs) and histone deacetylases (HDACs), and identified one of the HATs, HAC2, belonging to CBP family was required for TMV accumulation in Arabidopsis protoplasts. Combined these findings indicate that regulation of host gene expression, TF and histone modification are all involved in the early stage of virus-host interaction. Controlling these factors which are altered by virus infection in early stage may be a practicable and valid strategy in virus management. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-08T04:15:52Z (GMT). No. of bitstreams: 1 ntu-99-F93633004-1.pdf: 5841915 bytes, checksum: 281400fe4148acea53d8e7162777a1c2 (MD5) Previous issue date: 2010 | en |
| dc.description.tableofcontents | 誌謝 i
中文摘要 iii Abstract iv Chapter 1 : Introduction 1 1.1 Tobacco mosaic virus (TMV) 2 Taxonomy 2 Genome organization 2 1.2 TMV and model systems 3 1.3 Virus infection 5 Infection cycle of TMV 5 Virus-host interaction 7 Host cellular remodeling during plant virus infection 9 Viral and host proteins required for membrane targeting and virus replication complexes formation 11 1.4 Histone and gene regulation 13 Histone 13 Histone acetylation 14 Acetylation of non-histone targets 16 1.5 Antiviral Strategies 17 1.6 Early events of virus replication and the application in virus control 21 Aim in this thesis 22 Chapter 2 : Unusual targets identified for antiviral strategies : a systematic (screening) approach focused on early stages of virus replication 23 中文摘要 24 Abstract 25 Introduction 27 Materials and methods 31 Results 45 Discussion 51 Tables 55 Table 1. Primers used in this study 55 Table 2. Genes induced in common by TMV*CP.MP at 0.5, 4 and 6 hour 59 Table 3. Determination of interactions between PAP85 and P126 proteins by yeast two-hybrid assay 60 Figures 61 Fig. 1. Schematic representation of Tobacco mosaic virus (TMV) and derived constructs and detection of virus accumulation by northern blot hybridization 62 Fig. 2. Transfection efficiency analysis of TMV-infected Arabidopsis thaliana protoplasts 63 Fig. 3. Time course analysis of TMV*CP.MP accumulation in Arabidopsis protoplasts 64 Fig. 4. Venn diagram and in silico analysis of Arabidopsis genes up-regulated during TMV*CP.MP infection 65 Fig. 5. Confirmation of the relative expression of PAP85 obtained from microarray screening and the in silico gene expression of PAP85 by quantitative real-time RT-PCR 66 Fig. 6. Analysis of PAP85 expression in Arabidopsis protoplasts which were introduced in vitro-prepared PAP85 double-stranded RNA (dsRNA) 67 Fig. 7. Detection of virus accumulation in PAP85 dsRNA-treated protoplasts and quantification of the PAP85 RNA level in these cells 68 Fig. 8. Phenotype of wild-type and pap85-RNAi transgenic Arabidopsis 69 Fig. 9. The accumulation of TMV in pap85-RNAi transgenic Arabidopsis 70 Fig. 10. Complementation of PAP85 in Arabidopsis plants by Agrobacterium - mediated transient expression 71 Fig. 11. The effect of PAP85 on TMV forming the membrane-bound virus replication complexes (VRCs) 72 Fig. 12. Statistical analysis of the number of VRCs in Fig. 11 and quantification of PAP85 expression by real-time RT-PCR 73 Fig. 13. Subcellular localization of PAP85, TMV P126 and 11S globulin (At1g03880) in Arabidopsis protoplasts 75 Fig. 14. Analysis of interaction between PAP85 and TMV-P126 by BiFC 76 Fig. 15. The accumulation of TMV, CMV, ORSV and PVX in pap85 transgenic plants 77 Fig. 16. A systematic approach to identify gene(s) suitable for antiviral targets 78 Fig. 17. Relative expression of PAP85 in pCass2 vector and pCass2-P126 inoculated Arabidopsis protoplasts 79 Fig. 18. A model is illustrated to explain my results 80 Chapter 3 : Plant defense-related transcription factor WRKY6 plays both supportive and inhibitory roles in Tobacco mosaic virus infection 81 中文摘要 82 Abstract 83 Introduction 84 Materials and Methods 87 Results 91 Discussion 94 Tables 98 Table 1. Primers used in this study 98 Table 2. Genes with greater than 3-fold expression induced by Tobacco mosaic virus at 4 hr post-inoculation 99 Figures 100 Fig. 1. Relative expression of At1g62300 (WRKY6) in TMV*CP.MP-inoculated Arabidopsis protoplasts 100 Fig. 2. Phenotype of wild-type (WT), WRKY6-overexpressed (CaMV 35S::WRKY6) and -knockout (wrky6) plants, and the expression of WRKY6 in these plants 101 Fig. 3. The accumulation of TMV in WT, CaMV 35S::WRKY6 and wrky6 protoplasts 102 Fig. 4. The accumulation of TMV in WT, CaMV 35S::WRKY6 and wrky6 plants 103 Chapter 4 : Arabidopsis histone acetyltransferase, HAC2, involved in the accumulation of Tobacco mosaic virus in protoplasts 104 中文摘要 105 Abstract 106 Introduction 107 Material and Methods 109 Results 112 Discussion 113 Tables 115 Table 1. Selecting and screening mutant lines of Arabidopsis HAT and HDAC 115 Table 2. Primers used in this study 117 Figures 119 Fig. 1. Selection of homozygote lines of hac2 (SALK_049434) 119 Fig. 2. The accumulation of TMV in wild type and hac2 protoplasts 120 Conclusion 121 References 124 | |
| dc.language.iso | en | |
| dc.subject | 組蛋白修飾 | zh_TW |
| dc.subject | 菸草嵌紋病毒 | zh_TW |
| dc.subject | 阿拉伯芥 | zh_TW |
| dc.subject | PAP85 | zh_TW |
| dc.subject | WRKY6 | zh_TW |
| dc.subject | WRKY6 | en |
| dc.subject | histone modification | en |
| dc.subject | Tobacco mosaic virus | en |
| dc.subject | Arabidopsis thaliana | en |
| dc.subject | PAP85 | en |
| dc.title | 可做抗病毒目標之寄主因子篩選 | zh_TW |
| dc.title | Identification of host factors for antiviral targets | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 98-2 | |
| dc.description.degree | 博士 | |
| dc.contributor.oralexamcommittee | 蘇鴻基,吳素幸,鄭秋萍,洪挺軒 | |
| dc.subject.keyword | 菸草嵌紋病毒,阿拉伯芥,PAP85,WRKY6,組蛋白修飾, | zh_TW |
| dc.subject.keyword | Tobacco mosaic virus,Arabidopsis thaliana,PAP85,WRKY6,histone modification, | en |
| dc.relation.page | 140 | |
| dc.rights.note | 未授權 | |
| dc.date.accepted | 2010-08-05 | |
| dc.contributor.author-college | 生物資源暨農學院 | zh_TW |
| dc.contributor.author-dept | 植物病理與微生物學研究所 | zh_TW |
| 顯示於系所單位： | 植物病理與微生物學系 | |
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