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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
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dc.contributor.advisor | 林诗舜(Shih-Shun Lin) | |
dc.contributor.author | Pedro Arthuro Santa Cruz Peralta | en |
dc.contributor.author | 詹志宏 | zh_TW |
dc.date.accessioned | 2021-06-16T09:23:07Z | - |
dc.date.available | 2020-08-24 | |
dc.date.copyright | 2020-08-24 | |
dc.date.issued | 2020 | |
dc.date.submitted | 2020-08-14 | |
dc.identifier.citation | Alazem, M., He, M.-H., Moffett, P., and Lin, N.-S. (2017). Abscisic acid induces resistance against bamboo mosaic virus through Argonaute 2 and 3. Plant Physiol. 174, 339-355. Anandalakshmi, R., Pruss, G.J., Ge, X., Marathe, R., Mallory, A.C., Smith, T.H., and Vance, V.B. (1998). A viral suppressor of gene silencing in plants. Proc. Natl. Acad. Sci. U.S.A. 95, 13079-13084. Arribas-Hernández, L., Marchais, A., Poulsen, C., Haase, B., Hauptmann, J., Benes, V., Meister, G., and Brodersen, P. (2016). The slicer activity of ARGONAUTE1 is required specifically for the phasing, not production, of trans-acting short interfering RNAs in Arabidopsis. Plant Cell. 28, 1563-1580. Azevedo, J., Garcia, D., Pontier, D., Ohnesorge, S., Yu, A., Garcia, S., Braun, L., Bergdoll, M., Hakimi, M.A., Lagrange, T., and Voinnet, O. (2010). Argonaute quenching and global changes in Dicer homeostasis caused by a pathogen-encoded GW repeat protein. Genes Dev. 24, 904-915. Blevins, T., Rajeswaran, R., Shivaprasad, P.V., Beknazariants, D., Si-Ammour, A., Park, H.-S., Vazquez, F., Robertson, D., Meins Jr, F., and Hohn, T. (2006). Four plant Dicers mediate viral small RNA biogenesis and DNA virus induced silencing. Nucleic Acids Res. 34, 6233-6246. Bohmert, K., Camus, I., Bellini, C., Bouchez, D., Caboche, M., and Benning, C. (1998). AGO1 defines a novel locus of Arabidopsis controlling leaf development. EMBO J. 17, 170-180. Brodersen, P., Sakvarelidze-Achard, L., Bruun-Rasmussen, M., Dunoyer, P., Yamamoto, Y.Y., Sieburth, L., and Voinnet, O. (2008). Widespread translational inhibition by plant miRNAs and siRNAs. Science. 320, 1185-1190. Brosseau, C., and Moffett, P. (2015). Functional and genetic analysis identify a role for Arabidopsis ARGONAUTE5 in antiviral RNA silencing. Plant Cell 27, 1742-1754. Brosseau, C., El Oirdi, M., Adurogbangba, A., Ma, X., and Moffett, P. (2016). Antiviral defense involves AGO4 in an Arabidopsis–Potexvirus Interaction. Mol. Plant Microbe Interact. 29, 878-888. Brosseau, C., Bolaji, A., Roussin‐Léveillée, C., Zhao, Z., Biga, S., and Moffett, P. (2020). Natural variation in the Arabidopsis AGO2 gene is associated with susceptibility to potato virus X. New Phytol. 226, 866-878. Cao, Z., Zhang, J., Li, Y., Xu, X., Liu, G., Bhattacharrya, M.K., Yang, H., and Ren, D. (2007). Preparation of polyclonal antibody specific for AtPLC4, an Arabidopsis phosphatidylinositol-specific phospholipase C in rabbits. Protein Exp. Purif. 52, 306-312. Carrington, J.C., and Ambros, V. (2003). Role of microRNAs in plant and animal development. Science. 301, 336-338. Chiu, M.-T., Lin, C.-P., Lin, P.-C., and Lin, S.-S. (2013). Enhancement of IgG purification by FPLC for a serological study on the Turnip mosaic virus P1 protein. Plant Pathol. Bull. 22, 21-30. Chung, B.Y.-W., Valli, A., Deery, M.J., Navarro, F.J., Brown, K., Hnatova, S., Howard, J., Molnar, A., and Baulcombe, D.C. (2019). Distinct roles of Argonaute in the green alga Chlamydomonas reveal evolutionary conserved mode of miRNA-mediated gene expression. Sci. Rep. 9, 1-12. Curaba, J., and Chen, X. (2008). Biochemical activities of Arabidopsis RNA-dependent RNA polymerase 6. J. Biol. Chem. 283, 3059-3066. Deleris, A., Gallego-Bartolome, J., Bao, J., Kasschau, K.D., Carrington, J.C., and Voinnet, O. (2006). Hierarchical action and inhibition of plant Dicer-like proteins in antiviral defense. Science. 313, 68-71. Denli, A.M., and Hannon, G.J. (2003). RNAi: an ever-growing puzzle. Trends Biochem. Sci. 28, 196-201. Ding, S.-W., and Voinnet, O.J.C. (2007). Antiviral immunity directed by small RNAs 130, 413-426. Dunoyer, P., Schott, G., Himber, C., Meyer, D., Takeda, A., Carrington, J.C., and Voinnet, O. (2010). Small RNA duplexes function as mobile silencing signals between plant cells. Science. 328, 912-916. Dzianott, A., Sztuba-Solińska, J., and Bujarski, J.J. (2012). Mutations in the antiviral RNAi defense pathway modify Brome mosaic virus RNA recombinant profiles. Mol. Plant Microbe Interact. 25, 97-106. Fagard, M., Boutet, S., Morel, J.-B., Bellini, C., and Vaucheret, H. (2000). AGO1, QDE-2, and RDE-1 are related proteins required for post-transcriptional gene silencing in plants, quelling in fungi, and RNA interference in animals. Proc Natl Acad Sci U S A. 97, 11650-11654. Garcia-Ruiz, H., Takeda, A., Chapman, E.J., Sullivan, C.M., Fahlgren, N., Brempelis, K.J., and Carrington, J.C. (2010). Arabidopsis RNA-dependent RNA polymerases and dicer-like proteins in antiviral defense and small interfering RNA biogenesis during Turnip Mosaic Virus infection. Plant Cell. 22, 481-496. Garcia-Ruiz, H., Carbonell, A., Hoyer, J.S., Fahlgren, N., Gilbert, K.B., Takeda, A., Giampetruzzi, A., Ruiz, M.T.G., McGinn, M.G., and Lowery, N. (2015). Roles and programming of Arabidopsis ARGONAUTE proteins during Turnip mosaic virus infection. PLoS Pathog. 11, e1004755. Giner, A., Lakatos, L., García-Chapa, M., López-Moya, J.J., and Burgyán, J. (2010). Viral protein inhibits RISC activity by argonaute binding through conserved WG/GW motifs. PLoS Pathog. 6, e1000996. Harvey, J.J., Lewsey, M.G., Patel, K., Westwood, J., Heimstädt, S., Carr, J.P., and Baulcombe, D.C. (2011). An antiviral defense role of AGO2 in plants. PLoS One. 6, e14639. Hong, S.-F. (2015). Establishment of T-DNA Insertion Identification and Antibodies Affinity Purification for Studying the Mechanism on Autophagic AGO1 Degradation (unpublished Master Thesis: National Taiwan University). Hu, S.F., Wei, W.L., Hong, S.F., Fang, R.Y., Wu, H.Y., Lin, P.C., Sanobar, N., Wang, H.P., Sulistio, M., Wu, C.T., Lo, H.F., and Lin, S.S. (2020). Investigation of the effects of P1 on HC-Pro-mediated gene silencing suppression through genetics and omics approaches. Bot. Stud. 61. Incarbone, M., and Dunoyer, P. (2013). RNA silencing and its suppression: novel insights from in planta analyses. Trends Plant Sci. 18, 382-392. Jaubert, M., Bhattacharjee, S., Mello, A.F., Perry, K.L., and Moffett, P. (2011). ARGONAUTE2 mediates RNA-silencing antiviral defenses against Potato virus X in Arabidopsis. Plant Physiol. 156, 1556-1564. Jullien, P.E., Grob, S., Marchais, A., Pumplin, N., Chevalier, C., Bonnet, D.M., Otto, C., Schott, G., and Voinnet, O. (2018). Functional characterization of Arabidopsis ARGONAUTE 3 in reproductive tissues. Plant J. . Kasschau, K.D., and Carrington, J.C. (1998). A counterdefensive strategy of plant viruses: suppression of posttranscriptional gene silencing. Cell. 95, 461-470. Kasschau, K.D., Xie, Z., Allen, E., Llave, C., Chapman, E.J., Krizan, K.A., and Carrington, J.C. (2003). P1/HC-Pro, a viral suppressor of RNA silencing, interferes with Arabidopsis development and miRNA function. Genes Dev. 4, 205-217. Krieger, F., Möglich, A., and Kiefhaber, T. (2005). Effect of proline and glycine residues on dynamics and barriers of loop formation in polypeptide chains. J. Am. Chem. Soc. 127, 3346-3352. Kung, Y.-J., Lin, P.-C., Yeh, S.-D., Hong, S.-F., Chua, N.-H., Liu, L.-Y., Lin, C.-P., Huang, Y.-H., Wu, H.-W., and Chen, C.-C. (2014). Genetic analyses of the FRNK motif function of Turnip mosaic virus uncover multiple and potentially interactive pathways of cross-protection. Mol. Plant Microbe Interact. 27, 944-955. Lai, E.C. (2003). microRNAs: runts of the genome assert themselves. Curr. Biol. 13, R925-R936. Llave, C. (2010). Virus-derived small interfering RNAs at the core of plant–virus interactions. Trends Plant Sci. 15, 701-707. Llave, C., Xie, Z., Kasschau, K.D., and Carrington, J.C. (2002). Cleavage of Scarecrow-like mRNA targets directed by a class of Arabidopsis miRNA. Science. 297, 2053-2056. Ma, X., Nicole, M.-C., Meteignier, L.-V., Hong, N., Wang, G., and Moffett, P. (2015). Different roles for RNA silencing and RNA processing components in virus recovery and virus-induced gene silencing in plants. J Exp Bot. 66, 919-932. Mi, S., Cai, T., Hu, Y., Chen, Y., Hodges, E., Ni, F., Wu, L., Li, S., Zhou, H., and Long, C. (2008). Sorting of small RNAs into Arabidopsis argonaute complexes is directed by the 5′ terminal nucleotide. Cell. 133, 116-127. Morel, J.-B., Godon, C., Mourrain, P., Béclin, C., Boutet, S., Feuerbach, F., Proux, F., and Vaucheret, H. (2002). Fertile hypomorphic ARGONAUTE (ago1) mutants impaired in post-transcriptional gene silencing and virus resistance. Plant Cell. 14, 629-639. Qi, Y., Denli, A.M., and Hannon, G.J. (2005). Biochemical specialization within Arabidopsis RNA silencing pathways. Mol. Cell. 19, 421-428. Qi, Y., He, X., Wang, X.-J., Kohany, O., Jurka, J., and Hannon, G.J. (2006). Distinct catalytic and non-catalytic roles of ARGONAUTE4 in RNA-directed DNA methylation. Nature 443, 1008-1012. Qu, F. (2010). Antiviral role of plant-encoded RNA-dependent RNA polymerases revisited with deep sequencing of small interfering RNAs of virus origin. Mol. Plant Microbe Interact. 23, 1248-1252. Qu, F., Ye, X., and Morris, T.J. (2008). Arabidopsis DRB4, AGO1, AGO7, and RDR6 participate in a DCL4-initiated antiviral RNA silencing pathway negatively regulated by DCL1. Proc. Natl. Acad. Sci. U.S.A. 105, 14732-14737. Rodríguez-Leal, D., Castillo-Cobián, A., Rodríguez-Arévalo, I., and Vielle-Calzada, J.-P. (2016). A primary sequence analysis of the ARGONAUTE protein family in plants. Front. Plant Sci. 7, 1347. Sachetto-Martins, G., Franco, L.O., and de Oliveira, D.E. (2000). Plant glycine-rich proteins: a family or just proteins with a common motif? Biochim. Biophys. Acta. 1492, 1-14. Szittya, G., and Burgyán, J. (2013). RNA interference-mediated intrinsic antiviral immunity in plants. In Intrinsic immunity (Springer), pp. 153-181. Tabara, H., Sarkissian, M., Kelly, W.G., Fleenor, J., Grishok, A., Timmons, L., Fire, A., and Mello, C.C.J.C. (1999). The rde-1 gene, RNA interference, and transposon silencing in C. elegans 99, 123-132. Takeda, A., Iwasaki, S., Watanabe, T., Utsumi, M., and Watanabe, Y. (2008). The mechanism selecting the guide strand from small RNA duplexes is different among argonaute proteins. Plant Cell Physiol. 49, 493-500. Trier, N., Hansen, P., and Houen, G. (2019). Peptides, antibodies, peptide antibodies and more. Int. J. Mol. Sci. 20, 6289. Vaucheret, H.J.T.i.p.s. (2008). Plant argonautes 13, 350-358. Voinnet, O. (2008). Use, tolerance and avoidance of amplified RNA silencing by plants. Trends Plant Sci. 13, 317-328. Wang, J., Mei, J., and Ren, G. (2019). Plant microRNAs: biogenesis, homeostasis, and degradation. Front. Plant. Sci. 10, 360. Wang, X.-B., Jovel, J., Udomporn, P., Wang, Y., Wu, Q., Li, W.-X., Gasciolli, V., Vaucheret, H., and Ding, S.-W. (2011). The 21-nucleotide, but not 22-nucleotide, viral secondary small interfering RNAs direct potent antiviral defense by two cooperative argonautes in Arabidopsis thaliana. Plant Cell. 23, 1625-1638. Wu, H.-W., Lin, S.-S., Chen, K.-C., Yeh, S.-D., and Chua, N.-H. (2010). Discriminating mutations of HC-Pro of Zucchini yellow mosaic virus with differential effects on small RNA pathways involved in viral pathogenicity and symptom development. Mol. Plant Microbe Interact. 23, 17-28. Zhang, H., Xia, R., Meyers, B.C., and Walbot, V. (2015). Evolution, functions, and mysteries of plant ARGONAUTE proteins. Curr. Opin. Plant Biol. 27, 84-90. Zhang, X., Zhang, X., Singh, J., Li, D., and Qu, F. (2012). Temperature-dependent survival of Turnip crinkle virus-infected arabidopsis plants relies on an RNA silencing-based defense that requires dcl2, AGO2, and HEN1. J. Virol. 86, 6847-6854. Zhang, X., Yuan, Y.-R., Pei, Y., Lin, S.-S., Tuschl, T., Patel, D.J., and Chua, N.-H. (2006). Cucumber mosaic virus-encoded 2b suppressor inhibits Arabidopsis Argonaute1 cleavage activity to counter plant defense. Genes Dev. 20, 3255-3268. Zheng, X., Zhu, J., Kapoor, A., and Zhu, J.K. (2007). Role of Arabidopsis AGO6 in siRNA accumulation, DNA methylation and transcriptional gene silencing. EMBO J. 26, 1691-1701. | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/59417 | - |
dc.description.abstract | Argonaute (AGO) 蛋白是構成 RNA 誘導靜默複合體 (RISC) 的核心。 RISC 在植物中的作用之一是通過轉錄後基因靜默 (PTGS) 或轉錄基因靜默 (TGS) 介導抗病毒防禦。但是,某些植物病毒能夠通過表達RNA靜默病毒抑制子 (VSR) 來抑制宿主 PTGS與TGS,促進感染宿主,增加宿主易感性或出現症狀。另外,在一些研究中已經證明了AGO2在抗病毒防禦中的作用,包括對葉片中蕪菁嵌紋病毒 (TuMV) 的抗病毒作用。為了進一步闡明VSRs 對AGO累積的作用,我們製備 α-AGO2、α-AGO3 和 α-AGO10 IgG 抗體。AGOs N 端 (N) 具有高變異性,因此選擇此區域進行重組蛋白表達,以產出高專一性的 IgGs。我們發現 his-AGO2N 和 his-AGO3N 的高甘氨酸區干擾大腸桿菌重組蛋白表達。去除高甘氨酸區後,his-AGO2CoN 和 his-AGO3CoN (CoN; N 端區域的 C 末端) 可順利表達。由於 AGO3 和 AGO10 在阿拉伯芥中的低表達,α-AGO3 和 α-AGO10 沒辦法檢測到內源性蛋白AGO3 和 AGO10。但是,α-AGO2能檢測內源性蛋白 AGO2。Col-0,轉基因的阿拉伯芥株系,表現P1/HC-Pro 基因,是TuMV 的 VSR (P1/HC 植物) ,含有TuMV感染的Col-0被用來檢測AGO2 蛋白累積,我們觀察到P1/HC植物和感染 TuMV 的 Col-0 中 AGO2 蛋白積累減少。但 FPKM (Fragments Per Kilobase of transcript per Million mapped reads) 分析顯示 P1/HC 植物中 AGO2 轉錄子相較於 Col-0 的表現量上升。這些結果表明P1 /HC-Pro可能在轉譯後觸發 AGO2 降解,開啟了在植物與病毒相互作用的新研究方向。之後將進行進一步研究以了解 AGO2 在植物抗病毒反應中的作用。例如,利用受 TuMV 感染時其 AGO1 及 AGO2 小片段RNA (small RNA; sRNA) 圖譜確認及鑑定特定AGO的sRNAs。此外,通過免疫沉澱分析以AGO1或 AGO2為基礎的 RISC差異。 | zh_TW |
dc.description.abstract | Argonaute (AGO) proteins constitute the core of RNA-induced silencing complex (RISC). One of the roles of the RISC in plants is to mediate antiviral defense by post-transcriptional gene silencing (PTGS) or transcriptional gene silencing (TGS). However, plant viruses are able to inhibit host PTGS and TGS by expressing viral suppressors of RNA-silencing (VSRs) to facilitate their infection, resulting in an increase of host susceptibility or the development of symptoms. To further elucidate the effect of VSRs in AGO accumulation, we generated α-AGO2, α-AGO3, and α-AGO10 immunoglobulins G (IgGs). Notably, we used the N- terminus region (N) of AGOs for recombinant protein expression since it presents high variability, which would eventually allow the generation of IgGs with high specificity. Also, we found that a high-glycine rich region in the N-terminus of AGO2 and AGO3 interfered with his-AGO2N and his-AGO3N expression in E. coli. After the removal of the high-glycine regions, his-AGO2CoN and his-AGO3CoN (CoN; C-terminus end of the N-termini regions) could be expressed. Endogenous AGO3 and AGO10 proteins could not be detected by α-AGO3 and α-AGO10 perhaps due to their low expression levels in Arabidopsis. In contrast, endogenous AGO2 protein could be detected by α-AGO2. Arabidopsis Col-0, a transgenic Arabidopsis line expressing P1/HC-Pro, which is a VSR of turnip mosaic virus (TuMV) (P1/HC plants), and TuMV-infected Col-0 were used to investigate the accumulation of AGO2. We observed a decrease of AGO2 protein accumulation in P1/HC plants and TuMV-infected Col-0. However, the fragments per kilobase of transcript per million mapped reads (FPKM) value of AGO2 transcript showed upregulation in P1/HC plants compared to Col-0. These results suggest that P1/HC-Pro might trigger AGO2 reduction, which has novelty to open a new investigation direction for plant-virus interaction. Further studies such as sRNA profiling of IP AGO1 and AGO2 under TuMV infection will be used to identify and characterize AGO-specific sRNAs as well as AGO1 and AGO2 immunoprecipitation (IP) will be performed to elucidate the differences in AGO1-based and AGO2-based RISCs. | en |
dc.description.provenance | Made available in DSpace on 2021-06-16T09:23:07Z (GMT). No. of bitstreams: 1 U0001-1408202011032300.pdf: 4355998 bytes, checksum: 423937f4f5e13f4c2081698bcbd4c7b8 (MD5) Previous issue date: 2020 | en |
dc.description.tableofcontents | Contents Introduction 1 Materials and Methods 6 Plant materials and growth conditions 6 Infectious clone and mechanical inoculation 6 The his-AGO2CoN, his-AGO3CoN, his-AGO4N, his-AGO7N, his-AGO9N, and his-AGO10N synthesis into pET-28a 7 Recombinant protein purification by denatured-form protocol 8 Size-Exclusion Chromatography 9 Antibody production and purification 10 Western blot 11 IP assay and LC-MS/MS 11 AGO proteins FPKM analysis 13 Results 13 TuMV infection in Arabidopsis 13 Alignment of AGOs N-terminus regions 14 Cloning of codon-optimized AGOs DNA sequences 14 Secondary structure prediction of N-terminus AGOs 15 Recombinant proteins expression 16 α-AGO2 and α-AGO3 IgG tests 17 α-AGO10 IgG test 19 Endogenous AGO2 immunoprecipitation 20 AGOs expression in P1, HC and P1/HC transgenic plants 21 Discussion 22 A glycine-rich region might affect AGO peptide stability 22 The benefits of IgG production through recombinant protein inoculation approach 22 Low expression levels of endogenous proteins might affect homemade IgG detection ability 23 The P1/HC-Pro of TuMV might have an effect on AGO2 protein accumulation 24 Further studies on the roles of AGO1 and AGO2 in plant antiviral response 24 Conclusion 25 References 26 Tables and Figures 30 Supplementary Tables 48 | |
dc.language.iso | en | |
dc.title | Argonaute 抗體生產及 AGO2 在後轉錄時期基因靜默機制功能探討 | zh_TW |
dc.title | Generating Argonautes (AGOs) antibodies and investigating AGO2 functions in suppressing post-transcriptional gene silencing | en |
dc.type | Thesis | |
dc.date.schoolyear | 108-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 林劭品(Shau-Ping Lin),吳素幸(Shu-Hsing Wu) | |
dc.subject.keyword | 轉錄後基因靜默,蛋白,RNA靜默病毒抑制子, | zh_TW |
dc.subject.keyword | post-transcriptional gene silencing,argonaute,viral-suppressor of RNA-silencing, | en |
dc.relation.page | 50 | |
dc.identifier.doi | 10.6342/NTU202003381 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2020-08-17 | |
dc.contributor.author-college | 生物資源暨農學院 | zh_TW |
dc.contributor.author-dept | 生物科技研究所 | zh_TW |
顯示於系所單位: | 生物科技研究所 |
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