72 research outputs found
Roles and Programming of Arabidopsis ARGONAUTE Proteins during Turnip Mosaic Virus Infection
[EN] In eukaryotes, ARGONAUTE proteins (AGOs) associate with microRNAs (miRNAs), short interfering RNAs (siRNAs), and other classes of small RNAs to regulate target RNA or target loci. Viral infection in plants induces a potent and highly specific antiviral RNA silencing response characterized by the formation of virus-derived siRNAs. Arabidopsis thaliana has ten AGO genes of which AGO1, AGO2, and AGO7 have been shown to play roles in antiviral defense. A genetic analysis was used to identify and characterize the roles of AGO proteins in antiviral defense against Turnip mosaic virus (TuMV) in Arabidopsis. AGO1, AGO2 and AGO10 promoted anti-TuMV defense in a modular way in various organs, with AGO2 providing a prominent antiviral role in leaves. AGO5, AGO7 and AGO10 had minor effects in leaves. AGO1 and AGO10 had overlapping antiviral functions in inflorescence tissues after systemic movement of the virus, although the roles of AGO1 and AGO10 accounted for only a minor amount of the overall antiviral activity. By combining AGO protein immunoprecipitation with high-throughput sequencing of associated small RNAs, AGO2, AGO10, and to a lesser extent AGO1 were shown to associate with siRNAs derived from silencing suppressor (HC-Pro)-deficient TuMV-AS9, but not with siRNAs derived from wild-type TuMV. Co-immunoprecipitation and small RNA sequencing revealed that viral siRNAs broadly associated with wild-type HC-Pro during TuMV infection. These results support the hypothesis that suppression of antiviral silencing during TuMV infection, at least in part, occurs through sequestration of virus-derived siRNAs away from antiviral AGO proteins by HC-Pro. These findings indicate that distinct AGO proteins function as antiviral modules, and provide a molecular explanation for the silencing suppressor activity of HC-Pro.National Institutes of Health (www.nih.gov) grant AI43288 to JCC. National Science Foundation (www.nsf.gov) grant MCB-0956526 to JCC. Helen Hay Whitney (www.hhwf.org) Post-Doctoral fellowship (F-972) to HGR. USDA AFRI NIFA (www.csrees.usda.gov) Postdoctoral Fellowship (MOW-2012-01361) to NF. NSF (www.nsf.gov) Graduate Research Fellowship (DGE-1143954) to JSH Japan Society for the Promotion of Science (www.jsps.go.jp) Postdoctoral Fellowship to AT. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Garcia-Ruiz, H.; Carbonell, A.; Hoyer, JS.; Fahlgren, N.; Gilbert, KB.; Takeda, A.; Giampetruzzi, A.... (2015). Roles and Programming of Arabidopsis ARGONAUTE Proteins during Turnip Mosaic Virus Infection. PLoS Pathogens. 11(3). https://doi.org/10.1371/journal.ppat.1004755S113Ding, S.-W., & Voinnet, O. (2007). Antiviral Immunity Directed by Small RNAs. Cell, 130(3), 413-426. doi:10.1016/j.cell.2007.07.039Pumplin, N., & Voinnet, O. (2013). RNA silencing suppression by plant pathogens: defence, counter-defence and counter-counter-defence. Nature Reviews Microbiology, 11(11), 745-760. doi:10.1038/nrmicro3120Deleris, A., Gallego-Bartolome, J., Bao, J., Kasschau, K. D., Carrington, J. C., & Voinnet, O. (2006). Hierarchical Action and Inhibition of Plant Dicer-Like Proteins in Antiviral Defense. Science, 313(5783), 68-71. doi:10.1126/science.1128214Diaz-Pendon, J. A., Li, F., Li, W.-X., & Ding, S.-W. (2007). Suppression of Antiviral Silencing by Cucumber Mosaic Virus 2b Protein in Arabidopsis Is Associated with Drastically Reduced Accumulation of Three Classes of Viral Small Interfering RNAs. The Plant Cell, 19(6), 2053-2063. doi:10.1105/tpc.106.047449Curtin, S. J., Watson, J. M., Smith, N. A., Eamens, A. L., Blanchard, C. L., & Waterhouse, P. M. (2008). The roles of plant dsRNA-binding proteins in RNAi-like pathways. FEBS Letters, 582(18), 2753-2760. doi:10.1016/j.febslet.2008.07.004Qu, F., Ye, X., & Morris, T. J. (2008). Arabidopsis DRB4, AGO1, AGO7, and RDR6 participate in a DCL4-initiated antiviral RNA silencing pathway negatively regulated by DCL1. Proceedings of the National Academy of Sciences, 105(38), 14732-14737. doi:10.1073/pnas.0805760105Bologna, N. G., & Voinnet, O. (2014). The Diversity, Biogenesis, and Activities of Endogenous Silencing Small RNAs in Arabidopsis. Annual Review of Plant Biology, 65(1), 473-503. doi:10.1146/annurev-arplant-050213-035728Schuck, J., Gursinsky, T., Pantaleo, V., Burgyán, J., & Behrens, S.-E. (2013). AGO/RISC-mediated antiviral RNA silencing in a plant in vitro system. Nucleic Acids Research, 41(9), 5090-5103. doi:10.1093/nar/gkt193Brodersen, P., Sakvarelidze-Achard, L., Bruun-Rasmussen, M., Dunoyer, P., Yamamoto, Y. Y., Sieburth, L., & Voinnet, O. (2008). Widespread Translational Inhibition by Plant miRNAs and siRNAs. Science, 320(5880), 1185-1190. doi:10.1126/science.1159151Ciomperlik, J. J., Omarov, R. T., & Scholthof, H. B. (2011). An antiviral RISC isolated from Tobacco rattle virus-infected plants. Virology, 412(1), 117-124. doi:10.1016/j.virol.2010.12.018Iwakawa, H., & Tomari, Y. (2013). Molecular Insights into microRNA-Mediated Translational Repression in Plants. Molecular Cell, 52(4), 591-601. doi:10.1016/j.molcel.2013.10.033Huntzinger, E., & Izaurralde, E. (2011). Gene silencing by microRNAs: contributions of translational repression and mRNA decay. Nature Reviews Genetics, 12(2), 99-110. doi:10.1038/nrg2936Incarbone, M., & Dunoyer, P. (2013). RNA silencing and its suppression: novel insights from in planta analyses. Trends in Plant Science, 18(7), 382-392. doi:10.1016/j.tplants.2013.04.001Baumberger, N., Tsai, C.-H., Lie, M., Havecker, E., & Baulcombe, D. C. (2007). The Polerovirus Silencing Suppressor P0 Targets ARGONAUTE Proteins for Degradation. Current Biology, 17(18), 1609-1614. doi:10.1016/j.cub.2007.08.039Bortolamiol, D., Pazhouhandeh, M., Marrocco, K., Genschik, P., & Ziegler-Graff, V. (2007). The Polerovirus F Box Protein P0 Targets ARGONAUTE1 to Suppress RNA Silencing. Current Biology, 17(18), 1615-1621. doi:10.1016/j.cub.2007.07.061Csorba, T., Lózsa, R., Hutvágner, G., & Burgyán, J. (2010). Polerovirus protein P0 prevents the assembly of small RNA-containing RISC complexes and leads to degradation of ARGONAUTE1. The Plant Journal, 62(3), 463-472. doi:10.1111/j.1365-313x.2010.04163.xDerrien, B., Baumberger, N., Schepetilnikov, M., Viotti, C., De Cillia, J., Ziegler-Graff, V., … Genschik, P. (2012). Degradation of the antiviral component ARGONAUTE1 by the autophagy pathway. Proceedings of the National Academy of Sciences, 109(39), 15942-15946. doi:10.1073/pnas.1209487109Zhang, X., Yuan, Y.-R., Pei, Y., Lin, S.-S., Tuschl, T., Patel, D. J., & Chua, N.-H. (2006). Cucumber mosaic virus-encoded 2b suppressor inhibits Arabidopsis Argonaute1 cleavage activity to counter plant defense. Genes & Development, 20(23), 3255-3268. doi:10.1101/gad.1495506Giner, A., Lakatos, L., García-Chapa, M., López-Moya, J. J., & Burgyán, J. (2010). Viral Protein Inhibits RISC Activity by Argonaute Binding through Conserved WG/GW Motifs. PLoS Pathogens, 6(7), e1000996. doi:10.1371/journal.ppat.1000996Nakahara, K. S., & Masuta, C. (2014). Interaction between viral RNA silencing suppressors and host factors in plant immunity. Current Opinion in Plant Biology, 20, 88-95. doi:10.1016/j.pbi.2014.05.004Garcia-Ruiz, H., Takeda, A., Chapman, E. J., Sullivan, C. M., Fahlgren, N., Brempelis, K. J., & 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
. The Plant Cell, 22(2), 481-496. doi:10.1105/tpc.109.073056Vaucheret, H. (2008). Plant ARGONAUTES. Trends in Plant Science, 13(7), 350-358. doi:10.1016/j.tplants.2008.04.007Morel, J.-B., Godon, C., Mourrain, P., Béclin, C., Boutet, S., Feuerbach, F., … Vaucheret, H. (2002). Fertile Hypomorphic ARGONAUTE (ago1) Mutants Impaired in Post-Transcriptional Gene Silencing and Virus Resistance. The Plant Cell, 14(3), 629-639. doi:10.1105/tpc.010358Harvey, J. J. W., Lewsey, M. G., Patel, K., Westwood, J., Heimstädt, S., Carr, J. P., & Baulcombe, D. C. (2011). An Antiviral Defense Role of AGO2 in Plants. PLoS ONE, 6(1), e14639. doi:10.1371/journal.pone.0014639Zhang, X., Zhang, X., Singh, J., Li, D., & 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. Journal of Virology, 86(12), 6847-6854. doi:10.1128/jvi.00497-12Wang, X.-B., Jovel, J., Udomporn, P., Wang, Y., Wu, Q., Li, W.-X., … 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
. The Plant Cell, 23(4), 1625-1638. doi:10.1105/tpc.110.082305Dzianott, A., Sztuba-Solińska, J., & Bujarski, J. J. (2012). Mutations in the Antiviral RNAi Defense Pathway Modify Brome mosaic virus RNA Recombinant Profiles. Molecular Plant-Microbe Interactions®, 25(1), 97-106. doi:10.1094/mpmi-05-11-0137Carbonell, A., Fahlgren, N., Garcia-Ruiz, H., Gilbert, K. B., Montgomery, T. A., Nguyen, T., … Carrington, J. C. (2012). Functional Analysis of Three Arabidopsis ARGONAUTES Using Slicer-Defective Mutants
. The Plant Cell, 24(9), 3613-3629. doi:10.1105/tpc.112.099945Takeda, A., Iwasaki, S., Watanabe, T., Utsumi, M., & Watanabe, Y. (2008). The Mechanism Selecting the Guide Strand from Small RNA Duplexes is Different Among Argonaute Proteins. Plant and Cell Physiology, 49(4), 493-500. doi:10.1093/pcp/pcn043Azevedo, J., Garcia, D., Pontier, D., Ohnesorge, S., Yu, A., Garcia, S., … Voinnet, O. (2010). Argonaute quenching and global changes in Dicer homeostasis caused by a pathogen-encoded GW repeat protein. Genes & Development, 24(9), 904-915. doi:10.1101/gad.1908710Wei, W., Ba, Z., Gao, M., Wu, Y., Ma, Y., Amiard, S., … Qi, Y. (2012). A Role for Small RNAs in DNA Double-Strand Break Repair. Cell, 149(1), 101-112. doi:10.1016/j.cell.2012.03.002Zhang, X., Zhao, H., Gao, S., Wang, W.-C., Katiyar-Agarwal, S., Huang, H.-D., … Jin, H. (2011). Arabidopsis Argonaute 2 Regulates Innate Immunity via miRNA393∗-Mediated Silencing of a Golgi-Localized SNARE Gene, MEMB12. Molecular Cell, 42(3), 356-366. doi:10.1016/j.molcel.2011.04.010Zhu, H., Hu, F., Wang, R., Zhou, X., Sze, S.-H., Liou, L. W., … Zhang, X. (2011). Arabidopsis Argonaute10 Specifically Sequesters miR166/165 to Regulate Shoot Apical Meristem Development. Cell, 145(2), 242-256. doi:10.1016/j.cell.2011.03.024Mallory, A. C., Hinze, A., Tucker, M. R., Bouché, N., Gasciolli, V., Elmayan, T., … Laux, T. (2009). Redundant and Specific Roles of the ARGONAUTE Proteins AGO1 and ZLL in Development and Small RNA-Directed Gene Silencing. PLoS Genetics, 5(9), e1000646. doi:10.1371/journal.pgen.1000646Kasschau, K. D., Cronin, S., & Carrington, J. C. (1997). Genome Amplification and Long-Distance Movement Functions Associated with the Central Domain of Tobacco Etch Potyvirus Helper Component–Proteinase. Virology, 228(2), 251-262. doi:10.1006/viro.1996.8368Lakatos, L., Csorba, T., Pantaleo, V., Chapman, E. J., Carrington, J. C., Liu, Y.-P., … Burgyán, J. (2006). Small RNA binding is a common strategy to suppress RNA silencing by several viral suppressors. The EMBO Journal, 25(12), 2768-2780. doi:10.1038/sj.emboj.7601164Mallory, A. C., Reinhart, B. J., Bartel, D., Vance, V. B., & Bowman, L. H. (2002). A viral suppressor of RNA silencing differentially regulates the accumulation of short interfering RNAs and micro-RNAs in tobacco. Proceedings of the National Academy of Sciences, 99(23), 15228-15233. doi:10.1073/pnas.232434999Chapman, E. J. (2004). Viral RNA silencing suppressors inhibit the microRNA pathway at an intermediate step. Genes & Development, 18(10), 1179-1186. doi:10.1101/gad.1201204Kasschau, K. D., Xie, Z., Allen, E., Llave, C., Chapman, E. J., Krizan, K. A., & Carrington, J. C. (2003). P1/HC-Pro, a Viral Suppressor of RNA Silencing, Interferes with Arabidopsis Development and miRNA Function. Developmental Cell, 4(2), 205-217. doi:10.1016/s1534-5807(03)00025-xSchott, G., Mari-Ordonez, A., Himber, C., Alioua, A., Voinnet, O., & Dunoyer, P. (2012). Differential effects of viral silencing suppressors on siRNA and miRNA loading support the existence of two distinct cellular pools of ARGONAUTE1. The EMBO Journal, 31(11), 2553-2565. doi:10.1038/emboj.2012.92Shiboleth, Y. M., Haronsky, E., Leibman, D., Arazi, T., Wassenegger, M., Whitham, S. A., … Gal-On, A. (2007). The Conserved FRNK Box in HC-Pro, a Plant Viral Suppressor of Gene Silencing, Is Required for Small RNA Binding and Mediates Symptom Development. Journal of Virology, 81(23), 13135-13148. doi:10.1128/jvi.01031-07Endres, M. W., Gregory, B. D., Gao, Z., Foreman, A. W., Mlotshwa, S., Ge, X., … Vance, V. (2010). Two Plant Viral Suppressors of Silencing Require the Ethylene-Inducible Host Transcription Factor RAV2 to Block RNA Silencing. PLoS Pathogens, 6(1), e1000729. doi:10.1371/journal.ppat.1000729Ala-Poikela, M., Goytia, E., Haikonen, T., Rajamäki, M.-L., & Valkonen, J. P. T. (2011). Helper Component Proteinase of the Genus Potyvirus Is an Interaction Partner of Translation Initiation Factors eIF(iso)4E and eIF4E and Contains a 4E Binding Motif. Journal of Virology, 85(13), 6784-6794. doi:10.1128/jvi.00485-11Anandalakshmi, R., Marathe, R., Ge, X., Herr, J. M., Mau, C., Mallory, A., … Vance, V. B. (2000). A Calmodulin-Related Protein That Suppresses Posttranscriptional Gene Silencing in Plants. Science, 290(5489), 142-144. doi:10.1126/science.290.5489.142Iki, T., Yoshikawa, M., Nishikiori, M., Jaudal, M. C., Matsumoto-Yokoyama, E., Mitsuhara, I., … Ishikawa, M. (2010). In Vitro Assembly of Plant RNA-Induced Silencing Complexes Facilitated by Molecular Chaperone HSP90. Molecular Cell, 39(2), 282-291. doi:10.1016/j.molcel.2010.05.014Ballut, L., Drucker, M., Pugnière, M., Cambon, F., Blanc, S., Roquet, F., … Badaoui, S. (2005). HcPro, a multifunctional protein encoded by a plant RNA virus, targets the 20S proteasome and affects its enzymic activities. Journal of General Virology, 86(9), 2595-2603. doi:10.1099/vir.0.81107-0Soitamo, A. J., Jada, B., & Lehto, K. (2011). HC-Pro silencing suppressor significantly alters the gene expression profile in tobacco leaves and flowers. BMC Plant Biology, 11(1), 68. doi:10.1186/1471-2229-11-68Lellis, A. D., Kasschau, K. D., Whitham, S. A., & Carrington, J. C. (2002). Loss-of-Susceptibility Mutants of Arabidopsis thaliana Reveal an Essential Role for eIF(iso)4E during Potyvirus Infection. Current Biology, 12(12), 1046-1051. doi:10.1016/s0960-9822(02)00898-9Montgomery, T. A., Howell, M. D., Cuperus, J. T., Li, D., Hansen, J. E., Alexander, A. L., … Carrington, J. C. (2008). Specificity of ARGONAUTE7-miR390 Interaction and Dual Functionality in TAS3 Trans-Acting siRNA Formation. Cell, 133(1), 128-141. doi:10.1016/j.cell.2008.02.033Mi, S., Cai, T., Hu, Y., Chen, Y., Hodges, E., Ni, F., … Qi, Y. (2008). Sorting of Small RNAs into Arabidopsis Argonaute Complexes Is Directed by the 5′ Terminal Nucleotide. Cell, 133(1), 116-127. doi:10.1016/j.cell.2008.02.034Wang, H., Zhang, X., Liu, J., Kiba, T., Woo, J., Ojo, T., … Wang, X. (2011). Deep sequencing of small RNAs specifically associated with Arabidopsis AGO1 and AGO4 uncovers new AGO functions. The Plant Journal, 67(2), 292-304. doi:10.1111/j.1365-313x.2011.04594.xMérai, Z., Kerényi, Z., Kertész, S., Magna, M., Lakatos, L., & Silhavy, D. (2006). Double-Stranded RNA Binding May Be a General Plant RNA Viral Strategy To Suppress RNA Silencing. Journal of Virology, 80(12), 5747-5756. doi:10.1128/jvi.01963-05Cao, M., Du, P., Wang, X., Yu, Y.-Q., Qiu, Y.-H., Li, W., … Ding, S.-W. (2014). Virus infection triggers widespread silencing of host genes by a distinct class of endogenous siRNAs inArabidopsis. Proceedings of the National Academy of Sciences, 111(40), 14613-14618. doi:10.1073/pnas.1407131111Omarov, R. T., Ciomperlik, J. J., & Scholthof, H. B. (2007). RNAi-associated ssRNA-specific ribonucleases in Tombusvirus P19 mutant-infected plants and evidence for a discrete siRNA-containing effector complex. Proceedings of the National Academy of Sciences, 104(5), 1714-1719. doi:10.1073/pnas.0608117104Pantaleo, V., Szittya, G., & Burgyán, J. (2007). Molecular Bases of Viral RNA Targeting by Viral Small Interfering RNA-Programmed RISC. Journal of Virology, 81(8), 3797-3806. doi:10.1128/jvi.02383-06Schmid, M., Davison, T. S., Henz, S. R., Pape, U. J., Demar, M., Vingron, M., … Lohmann, J. U. (2005). A gene expression map of Arabidopsis thaliana development. Nature Genetics, 37(5), 501-506. doi:10.1038/ng1543Baumberger, N., & Baulcombe, D. C. (2005). Arabidopsis ARGONAUTE1 is an RNA Slicer that selectively recruits microRNAs and short interfering RNAs. Proceedings of the National Academy of Sciences, 102(33), 11928-11933. doi:10.1073/pnas.0505461102Donaire, L., Barajas, D., Martínez-García, B., Martínez-Priego, L., Pagán, I., & Llave, C. (2008). Structural and Genetic Requirements for the Biogenesis of
Tobacco Rattle Virus
-Derived Small Interfering RNAs. Journal of Virology, 82(11), 5167-5177. doi:10.1128/jvi.00272-08Cao, M., Ye, X., Willie, K., Lin, J., Zhang, X., Redinbaugh, M. G., … Qu, F. (2010). The Capsid Protein of
Turnip Crinkle Virus
Overcomes Two Separate Defense Barriers To Facilitate Systemic Movement of the Virus in
Arabidopsis. Journal of Virology, 84(15), 7793-7802. doi:10.1128/jvi.02643-09Blevins, T., Rajeswaran, R., Shivaprasad, P. V., Beknazariants, D., Si-Ammour, A., Park, H.-S., … Pooggin, M. M. (2006). Four plant Dicers mediate viral small RNA biogenesis and DNA virus induced silencing. Nucleic Acids Research, 34(21), 6233-6246. doi:10.1093/nar/gkl886Wang, N., Zhang, D., Wang, Z., Xun, H., Ma, J., Wang, H., … Liu, B. (2014). Mutation of the RDR1 gene caused genome-wide changes in gene expression, regional variation in small RNA clusters and localized alteration in DNA methylation in rice. BMC Plant Biology, 14(1). doi:10.1186/1471-2229-14-177Montgomery, T. A., Yoo, S. J., Fahlgren, N., Gilbert, S. D., Howell, M. D., Sullivan, C. M., … Carrington, J. C. (2008). AGO1-miR173 complex initiates phased siRNA formation in plants. Proceedings of the National Academy of Sciences, 105(51), 20055-20062. doi:10.1073/pnas.0810241105Cuperus, J. T., Carbonell, A., Fahlgren, N., Garcia-Ruiz, H., Burke, R. T., Takeda, A., … Carrington, J. C. (2010). Unique functionality of 22-nt miRNAs in triggering RDR6-dependent siRNA biogenesis from target transcripts in Arabidopsis. Nature Structural & Molecular Biology, 17(8), 997-1003. doi:10.1038/nsmb.1866Rajeswaran, R., Aregger, M., Zvereva, A. S., Borah, B. K., Gubaeva, E. G., & Pooggin, M. M. (2012). Sequencing of RDR6-dependent double-stranded RNAs reveals novel features of plant siRNA biogenesis. Nucleic Acids Research, 40(13), 6241-6254. doi:10.1093/nar/gks242Chen, H.-M., Chen, L.-T., Patel, K., Li, Y.-H., Baulcombe, D. C., & Wu, S.-H. (2010). 22-nucleotide RNAs trigger secondary siRNA biogenesis in plants. Proceedings of the National Academy of Sciences, 107(34), 15269-15274. doi:10.1073/pnas.1001738107Allen, E., Xie, Z., Gustafson, A. M., & Carrington, J. C. (2005). microRNA-Directed Phasing during Trans-Acting siRNA Biogenesis in Plants. Cell, 121(2), 207-221. doi:10.1016/j.cell.2005.04.004Bhattacharjee, S., Zamora, A., Azhar, M. T., Sacco, M. A., Lambert, L. H., & Moffett, P. (2009). Virus resistance induced by NB-LRR proteins involves Argonaute4-dependent translational control. The Plant Journal, 58(6), 940-951. doi:10.1111/j.1365-313x.2009.03832.xCurtis, M. D., & Grossniklaus, U. (2003). A Gateway Cloning Vector Set for High-Throughput Functional Analysis of Genes in Planta. Plant Physiology, 133(2), 462-469. doi:10.1104/pp.103.027979Lobbes, D., Rallapalli, G., Schmidt, D. D., Martin, C., & Clarke, J. (2006). SERRATE: a new player on the plant microRNA scene. EMBO reports, 7(10), 1052-1058. doi:10.1038/sj.embor.7400806Agorio, A., & Vera, P. (2007). ARGONAUTE4 Is Required for Resistance to Pseudomonas syringae in Arabidopsis. The Plant Cell, 19(11), 3778-3790. doi:10.1105/tpc.107.054494Hunter, C., Sun, H., & Poethig, R. S. (2003). The Arabidopsis Heterochronic Gene ZIPPY Is an ARGONAUTE Family Member. Current Biology, 13(19), 1734-1739. doi:10.1016/j.cub.2003.09.004Kleinboelting, N., Huep, G., Kloetgen, A., Viehoever, P., & Weisshaar, B. (2011). GABI-Kat SimpleSearch: new features of the Arabidopsis thaliana T-DNA mutant database. Nucleic Acids Research, 40(D1), D1211-D1215. doi:10.1093/nar/gkr1047Alonso, J. M., Stepanova, A. N., Leisse, T. J., Kim, C. J., Chen, H., Shinn, P., … Ecker, J. R. (2003). Genome-Wide Insertional Mutagenesis of
Arabidopsis thaliana. Science, 301(5633), 653-657. doi:10.1126/science.1086391Clough, S. J., & Bent, A. F. (1998). Floral dip: a simplified method forAgrobacterium-mediated transformation ofArabidopsis thaliana. The Plant Journal, 16(6), 735-743. doi:10.1046/j.1365-313x.1998.00343.xVárallyay, É., Válóczi, A., Ágyi, Á., Burgyán, J., & Havelda, Z. (2010). Plant virus-mediated induction of miR168 is associated with repression of ARGONAUTE1 accumulation. The EMBO Journal, 29(20), 3507-3519. doi:10.1038/emboj.2010.215Fahlgren, N., Sullivan, C. M., Kasschau, K. D., Chapman, E. J., Cumbie, J. S., Montgomery, T. A., … Carrington, J. C. (2009). Computational and analytical framework for small RNA profiling by high-throughput sequencing. RNA, 15(5), 992-1002. doi:10.1261/rna.147380
Roles and Programming of Arabidopsis ARGONAUTE Proteins During Turnip Mosaic Virus Infection
In eukaryotes, ARGONAUTE proteins (AGOs) associate with microRNAs (miRNAs), short interfering RNAs (siRNAs), and other classes of small RNAs to regulate target RNA or target loci. Viral infection in plants induces a potent and highly specific antiviral RNA silencing response characterized by the formation of virus-derived siRNAs. Arabidopsis thaliana has ten AGO genes of which AGO1, AGO2, and AGO7 have been shown to play roles in antiviral defense. A genetic analysis was used to identify and characterize the roles of AGO proteins in antiviral defense against Turnip mosaic virus (TuMV) in Arabidopsis. AGO1, AGO2 and AGO10 promoted anti-TuMV defense in a modular way in various organs, with AGO2 providing a prominent antiviral role in leaves. AGO5, AGO7 and AGO10 had minor effects in leaves. AGO1 and AGO10 had overlapping antiviral functions in inflorescence tissues after systemic movement of the virus, although the roles of AGO1 and AGO10 accounted for only a minor amount of the overall antiviral activity. By combining AGO protein immunoprecipitation with high-throughput sequencing of associated small RNAs, AGO2, AGO10, and to a lesser extent AGO1 were shown to associate with siRNAs derived from silencing suppressor (HC-Pro)-deficient TuMV-AS9, but not with siRNAs derived from wild-type TuMV. Co-immunoprecipitation and small RNA sequencing revealed that viral siRNAs broadly associated with wild-type HC-Pro during TuMV infection. These results support the hypothesis that suppression of antiviral silencing during TuMV infection, at least in part, occurs through sequestration of virus-derived siRNAs away from antiviral AGO proteins by HC-Pro. These findings indicate that distinct AGO proteins function as antiviral modules, and provide a molecular explanation for the silencing suppressor activity of HC-Pro
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Phytophthora Have Distinct Endogenous Small RNA Populations That Include Short Interfering and microRNAs
In eukaryotes, RNA silencing pathways utilize 20-30-nucleotide small RNAs to regulate gene expression, specify and maintain chromatin structure, and repress viruses and mobile genetic elements. RNA silencing was likely present in the common ancestor of modern eukaryotes, but most research has focused on plant and animal RNA silencing systems. Phytophthora species belong to a phylogenetically distinct group of economically important plant pathogens that cause billions of dollars in yield losses annually as well as ecologically devastating outbreaks. We analyzed the small RNA-generating components of the genomes of P. infestans, P. sojae and P. ramorum using bioinformatics, genetic, phylogenetic and high-throughput sequencing-based methods. Each species produces two distinct populations of small RNAs that are predominantly 21- or 25-nucleotides long. The 25-nucleotide small RNAs were primarily derived from loci encoding transposable elements and we propose that these small RNAs define a pathway of short-interfering RNAs that silence repetitive genetic elements. The 21-nucleotide small RNAs were primarily derived from inverted repeats, including a novel microRNA family that is conserved among the three species, and several gene families, including Crinkler effectors and type III fibronectins. The Phytophthora microRNA is predicted to target a family of amino acid/auxin permeases, and we propose that 21-nucleotide small RNAs function at the post-transcriptional level. The functional significance of microRNA-guided regulation of amino acid/auxin permeases and the association of 21-nucleotide small RNAs with Crinkler effectors remains unclear, but this work provides a framework for testing the role of small RNAs in Phytophthora biology and pathogenesis in future work
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Roles and Programming of Arabidopsis ARGONAUTE Proteins during Turnip Mosaic Virus Infection
In eukaryotes, ARGONAUTE proteins (AGOs) associate with microRNAs (miRNAs), short interfering RNAs (siRNAs), and other classes of small RNAs to regulate target RNA or target loci. Viral infection in plants induces a potent and highly specific antiviral RNA silencing response characterized by the formation of virus-derived siRNAs. Arabidopsis thaliana has ten AGO genes of which AGO1, AGO2, and AGO7 have been shown to play roles in antiviral defense. A genetic analysis was used to identify and characterize the roles of AGO proteins in antiviral defense against Turnip mosaic virus (TuMV) in Arabidopsis. AGO1, AGO2 and AGO10 promoted anti-TuMV defense in a modular way in various organs, with AGO2 providing a prominent antiviral role in leaves. AGO5, AGO7 and AGO10 had minor effects in leaves. AGO1 and AGO10 had overlapping antiviral functions in inflorescence tissues after systemic movement of the virus, although the roles of AGO1 and AGO10 accounted for only a minor amount of the overall antiviral activity. By combining AGO protein immunoprecipitation with high-throughput sequencing of associated small RNAs, AGO2, AGO10, and to a lesser extent AGO1 were shown to associate with siRNAs derived from silencing suppressor (HC-Pro)-deficient TuMV-AS9, but not with siRNAs derived from wild-type TuMV. Co-immunoprecipitation and small RNA sequencing revealed that viral siRNAs broadly associated with wild-type HC-Pro during TuMV infection. These results support the hypothesis that suppression of antiviral silencing during TuMV infection, at least in part, occurs through sequestration of virus-derived siRNAs away from antiviral AGO proteins by HC-Pro. These findings indicate that distinct AGO proteins function as antiviral modules, and provide a molecular explanation for the silencing suppressor activity of HC-Pro
Intracerebroventricular Streptozotocin Injections as a Model of Alzheimer’s Disease: in Search of a Relevant Mechanism
The ε3 and ε4 Alleles of Human APOE Differentially Affect Tau Phosphorylation in Hyperinsulinemic and Pioglitazone Treated Mice
Impaired insulin signalling is increasingly thought to contribute to Alzheimer's disease (AD). The ε4 isoform of the APOE gene is the greatest genetic risk factor for sporadic, late onset AD, and is also associated with risk for type 2 diabetes mellitus (T2DM). Neuropathological studies reported the highest number of AD lesions in brain tissue of ε4 diabetic patients. However other studies assessing AD pathology amongst the diabetic population have produced conflicting reports and have failed to show an increase in AD-related pathology in diabetic brain. The thiazolidinediones (TZDs), peroxisome proliferator-activated receptor gamma agonists, are peripheral insulin sensitisers used to treat T2DM. The TZD, pioglitazone, improved memory and cognitive functions in mild to moderate AD patients. Since it is not yet clear how apoE isoforms influence the development of T2DM and its progression to AD, we investigated amyloid beta and tau pathology in APOE knockout mice, carrying human APOEε3 or ε4 transgenes after diet-induced insulin resistance with and without pioglitazone treatment.Male APOE knockout, APOEε3-transgenic and APOEε4-transgenic mice, together with background strain C57BL6 mice were kept on a high fat diet (HFD) or low fat diet (LFD) for 32 weeks, or were all fed HFD for 32 weeks and during the final 3 weeks animals were treated with pioglitazone or vehicle.All HFD animals developed hyperglycaemia with elevated plasma insulin. Tau phosphorylation was reduced at 3 epitopes (Ser396, Ser202/Thr205 and Thr231) in all HFD, compared to LFD, animals independent of APOE genotype. The introduction of pioglitazone to HFD animals led to a significant reduction in tau phosphorylation at the Ser202/Thr205 epitope in APOEε3 animals only. We found no changes in APP processing however the levels of soluble amyloid beta 40 was reduced in APOE knockout animals treated with pioglitazone
In vitro characterization of the chaperone activity of the Arabidopsis non-canonical small heat shock protein BOBBER
Small heat shock proteins (sHSP) are a diverse family of molecular chaperones which bind thermally denatured proteins and thereby prevent them from aggregating. This prevents damage to cells and facilitates their later refolding. Plant sHSP have been characterized biochemically, but their physiological importance is unclear because sHSP knock-out mutants have shown few phenotypes when subjected to heat stress, possibly because of redundancy.\ud
BOBBER1 (BOB) is 34.5 kDa Arabidopsis thaliana protein with homology to sHSPs that is induced by high temperatures and localizes to heat shock granules along with other sHSPs. bob1-3 mutants have thermotolerance defects and developmental irregularities, suggesting a novel role for BOB in the plant chaperone network.\ud
In this investigation, a recombinant 6xHis-tagged form of BOB protein was expressed in E. coli and purified. The ATP-independent chaperone activity of BOB was demonstrated; BOB was shown to reduce the thermal aggregation of the model substrates malate dehydrogenase and citrate synthase. Site-directed mutagenesis was used to create mutant alleles of BOB protein that were both computationally predicted to disrupt its structure and are available in plants.\ud
Truncated forms of the BOB N and C termini were also expressed and purified in an attempt to understand the roles of the protein domains in interaction with partially unfolded substrate. The N terminus did not reduce aggregation under any condition tested, while two different forms of the C termini showed reduced chaperone activity compared to full length protein.\ud
The point mutation found in thermotolerance defective bob1-3 mutants did not result in loss of chaperone activity, raising the possibility that the mutation disrupts interactions with specific substrate(s) or cochaperones which would not be reflected in our assay. Identification of these substrates may be required for an understanding of why BOB is necessary for normal development and thermotolerance
Time-lapse photograph dataset: complementation of ago7 mutant A. thaliana plants with truncated promoter transgenes
<p>This transgenic complementation experiment is described in a manuscript by Hoyer et al.</p>
<p>Time-stamped photographs are provided in twelve directories by camera (twelve overlapping fields of view) and a manifest file with file SHA-1 sums is included.</p>
<p>Metadata will be made available separately, to facilitate updates. Seed was plated and growth started on 2016-08-26.</p
Scans of leaves dissected in phyllotactic order: complementation of ago7 mutant A. thaliana plants with truncated promoter transgenes
<p>This transgenic complementation experiment is described in a manuscript by Hoyer et al.</p>
<p>Adaxial (ad) and abaxial (ab) surfaces of rosettes were photographed in between removal of leaves, to enable checking that leaves were removed in the correct phyllotactic order.<strong> </strong>A manifest file with file SHA-1 sums is included.</p>
<p>Metadata and LeafJ measurements will be made available separately, to facilitate updates. Seed was plated and growth started on 2016-08-26. Leaves were dissected and scanned 33 and 35 days post-stratification (2016-09-28 and 30).</p
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