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Expression of the Beet necrotic yellow vein virus p25 protein induces hormonal changes and a root branching phenotype in Arabidopsis thaliana

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Abstract

The RNA-3-encoded p25 protein was previously characterized as one of the major symptom determinants of the Beet necrotic yellow vein virus. Previous analyses reported the influence of the p25 protein in root proliferation phenotype observed in rhizomania disease on infected sugar beets (Beta vulgaris). A transgenic approach was developed, in which the p25 protein was constitutively expressed in Arabidopsis thaliana Columbia (Col-0) ecotype in order to provide new clues as to how the p25 protein might promote alone disease development and symptom expression. Transgenic plants were characterized by Southern blot and independent lines carrying single and multiple copies of the transgene were selected. Mapping of the T-DNA insertion was performed on the monocopy homozygote lines. P25 protein was localized both in the nucleus and in the cytoplasm of epidermal and root cells of transgenic plants. Although A. thaliana was not described as a susceptible host for BNYVV infection, abnormal root branching was observed on p25 protein-expressing A. thaliana plants. Moreover, these transgenic plants were more susceptible than wild-type plants to auxin analog treatment (2,4-D) but more resistant to methyl jasmonate (MeJA), abscisic acid (ABA) and to lesser extend to salicylic acid (SA). Hormonal content assays measuring plant levels of auxin (IAA), jasmonate (JA) and ethylene precursor (ACC) revealed major hormonal changes. Global transcript profiling analyses on roots displayed differential gene expressions that could corroborate root branching phenotype and stress signaling modifications.

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References

  • Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402

    Article  PubMed  CAS  Google Scholar 

  • Berardini TZ, Mundodi S, Reiser L, Huala E, Garcia-Hernandez M, Zhang P, Mueller LA, Yoon J, Doyle A, Lander G, Moseyko N, Yoo D, Xu I, Zoeckler B, Montoya M, Miller N, Weems D, Rhee SY (2004) Functional annotation of the Arabidopsis genome using controlled vocabularies. Plant Physiol 135:745–755

    Article  PubMed  CAS  Google Scholar 

  • Bialek K, Cohen JD (1989) Quantitation of indoleacetic acid conjugates in bean seeds by direct tissue hydrolysis. Plant Physiol 90:398–400

    Article  PubMed  CAS  Google Scholar 

  • Boyes DC, Zayed AM, Ascenzi R, McCaskill AJ, Hoffman NE, Davis KR, Gorlach J (2001) Growth stage-based phenotypic analysis of Arabidopsis: a model for high throughput functional genomics in plants. Plant Cell 13:1499–1510

    Article  PubMed  CAS  Google Scholar 

  • Bülow L, Schindler M, Hehl R (2007) PathoPlant®: a platform for microarray expression data to analyze co-regulated genes involved in plant defense responses. Nucleic Acids Res 35:D841–845

    Article  PubMed  Google Scholar 

  • Canet JV, Dobon A, Ibanez F, Perales L, Tornero P (2010) Resistance and biomass in Arabidopsis: a new model for salicylic acid perception. Plant Biotechnol J 8:126–141

    Article  PubMed  CAS  Google Scholar 

  • Chauvaux N, Van Dongen W, Esmans EL, Van Onckelen HA (1993) Liquid chromatographic-mass spectrometric determination of 1-aminocyclopropane-1-carboxylic acid in tobacco. J Chromatogr A 657:337–343

    Article  CAS  Google Scholar 

  • Chiba S, Miyanishi M, Andika IB, Kondo H, Tamada T (2008) Identification of amino acids of the beet necrotic yellow vein virus p25 protein required for induction of the resistance response in leaves of Beta vulgaris plants. J Gen Virol 89:1314–1323

    Article  PubMed  CAS  Google Scholar 

  • Cho H-T, Cosgrove DJ (2002) Regulation of root hair initiation and expansin gene expression in arabidopsis. Plant Cell 14:3237–3253

    Article  PubMed  CAS  Google Scholar 

  • Clough SJ, Bent AF (1998) Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J 16:735–743

    Article  PubMed  CAS  Google Scholar 

  • Corbineau F, Rudnicki RM, Come D (1988) The effect of methyl jasmonate on sunflower (Helianthus annuus L.) seed germination and seedling development. Plant Growth Regul 7:157–169

    CAS  Google Scholar 

  • Crowe ML, Serizet C, Thareau V, Aubourg S, Rouze P, Hilson P, Beynon J, Weisbeek P, van Hummelen P, Reymond P, Paz-Ares J, Nietfeld W, Trick M (2003) CATMA: a complete Arabidopsis GST database. Nucleic Acids Res 31:156–158

    Article  PubMed  CAS  Google Scholar 

  • Dunoyer P, Lecellier CH, Parizotto EA, Himber C, Voinnet O (2004) Probing the microRNA and small interfering RNA pathways with virus-encoded suppressors of RNA silencing. Plant Cell 16:1235–1250

    Article  PubMed  CAS  Google Scholar 

  • Falk A, Feys BJ, Frost LN, Jones JD, Daniels MJ, Parker JE (1999) EDS1, an essential component of R gene-mediated disease resistance in Arabidopsis has homology to eukaryotic lipases. Proc Natl Acad Sci USA 96:3292–3297

    Article  PubMed  CAS  Google Scholar 

  • Finkelstein R (1994) Mutations at two new Arabidospis ABA response loci are similar to the abi3 mutations. Plant J 5:765–771

    Article  Google Scholar 

  • Gagnot S, Tamby JP, Martin-Magniette ML, Bitton F, Taconnat L, Balzergue S, Aubourg S, Renou JP, Lecharny A, Brunaud V (2008) CATdb: a public access to Arabidopsis transcriptome data from the URGV-CATMA platform. Nucleic Acids Res 36:D986–D990

    Article  PubMed  CAS  Google Scholar 

  • Ge Y, Dudoit S, Speed TP (2003) Resampling-based testing for microarray data analysis. Test 12

  • Haeberle AM, Stussi-Garaud C (1995) In situ localization of the non-structural protein P25 encoded by beet necrotic yellow vein virus RNA 3. J Gen Virol 76:643–650

    Article  PubMed  CAS  Google Scholar 

  • Hand DJ, Taylor CC (1987) Multivariate analysis of variance and repeated measures. Chapman and Hall/CRC, London

    Google Scholar 

  • Hilson P, Allemeersch J, Altmann T, Aubourg S, Avon A, Beynon J, Bhalerao RP, Bitton F, Caboche M, Cannoot B, Chardakov V, Cognet-Holliger C, Colot V, Crowe M, Darimont C, Durinck S, Eickhoff H, de Longevialle AF, Farmer EE, Grant M, Kuiper MT, Lehrach H, Leon C, Leyva A, Lundeberg J, Lurin C, Moreau Y, Nietfeld W, Paz-Ares J, Reymond P, Rouze P, Sandberg G, Segura MD, Serizet C, Tabrett A, Taconnat L, Thareau V, Van Hummelen P, Vercruysse S, Vuylsteke M, Weingartner M, Weisbeek PJ, Wirta V, Wittink FR, Zabeau M, Small I (2004) Versatile gene-specific sequence tags for Arabidopsis functional genomics: transcript profiling and reverse genetics applications. Genome Res 14:2176–2189

    Article  PubMed  CAS  Google Scholar 

  • Himber C, Dunoyer P, Moissiard G, Ritzenthaler C, Voinnet O (2003) Transitivity-dependent and -independent cell-to-cell movement of RNA silencing. EMBO J 22:4523–4533

    Article  PubMed  CAS  Google Scholar 

  • Holsters M, Silva B, Van Vliet F, Genetello C, De Block M, Dhaese P, Depicker A, Inze D, Engler G, Villarroel R et al (1980) The functional organization of the nopaline A. tumefaciens plasmid pTiC58. Plasmid 3:212–230

    Article  PubMed  CAS  Google Scholar 

  • Huckelhoven R, Dechert C, Kogel KH (2003) Overexpression of barley BAX inhibitor 1 induces breakdown of mlo-mediated penetration resistance to Blumeria graminis. Proc Natl Acad Sci USA 100:5555–5560

    Article  PubMed  CAS  Google Scholar 

  • Ishikawa M, Naito S, Ohno T (1993) Effects of the tom1 mutation of Arabidopsis thaliana on the multiplication of tobacco mosaic virus RNA in protoplasts. J Virol 67:5328–5338

    PubMed  CAS  Google Scholar 

  • Jupin I, Guilley H, Richards KE, Jonard G (1992) Two proteins encoded by beet necrotic yellow vein virus RNA 3 influence symptom phenotype on leaves. EMBO J 11:479–488

    PubMed  CAS  Google Scholar 

  • Kawaoka A, Matsunaga E, Endo S, Kondo S, Yoshida K, Shinmyo A, Ebinuma H (2003) Ectopic expression of a horseradish peroxidase enhances growth rate and increases oxidative stress resistance in hybrid aspen. Plant Physiol 132:1177–1185

    Article  PubMed  CAS  Google Scholar 

  • Klein E, Link D, Schirmer A, Erhardt M, Gilmer D (2007) Sequence variation within Beet necrotic yellow vein virus p25 protein influences its oligomerization and isolate pathogenicity on Tetragonia expansa. Virus Res 126:53–61

    Article  PubMed  CAS  Google Scholar 

  • Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685

    Article  PubMed  CAS  Google Scholar 

  • Larson RL, Wintermantel WM, Hill A, Fortis L, Nunez A (2008) Proteome changes in sugar beet in response to Beet necrotic yellow vein virus. Physiol Mol Plant Pathol 72:62–72

    Article  CAS  Google Scholar 

  • Lauber E, Guilley H, Tamada T, Richards KE, Jonard G (1998) Vascular movement of beet necrotic yellow vein virus in Beta macrocarpa is probably dependent on an RNA 3 sequence domain rather than a gene product. J Gen Virol 79:385–393

    PubMed  CAS  Google Scholar 

  • Lurin C, Andres C, Aubourg S, Bellaoui M, Bitton F, Bruyere C, Caboche M, Debast C, Gualberto J, Hoffmann B, Lecharny A, Le Ret M, Martin-Magniette ML, Mireau H, Peeters N, Renou JP, Szurek B, Taconnat L, Small I (2004) Genome-wide analysis of Arabidopsis pentatricopeptide repeat proteins reveals their essential role in organelle biogenesis. Plant Cell 16:2089–2103

    Article  PubMed  CAS  Google Scholar 

  • Netting AG, Milborrow BV (1988) Methane chemical ionization Mass-Spectrometry of the pentafluorobenzyl derivatives of Abscisic-acid, its metabolites and other plant-growth regulators. Biomed Environ Mass Spectrom 17:281–286

    Article  CAS  Google Scholar 

  • Niesbach-Klosgen U, Guilley H, Jonard G, Richards K (1990) Immunodetection in vivo of beet necrotic yellow vein virus-encoded proteins. Virology 178:52–61

    Article  PubMed  CAS  Google Scholar 

  • Ottenschlager I, Wolff P, Wolverton C, Bhalerao RP, Sandberg G, Ishikawa H, Evans M, Palme K (2003) Gravity-regulated differential auxin transport from columella to lateral root cap cells. Proc Natl Acad Sci USA 100:2987–2991

    Article  PubMed  CAS  Google Scholar 

  • Pan X, Welti R, Wang X (2008) Simultaneous quantification of major phytohormones and related compounds in crude plant extracts by liquid chromatography-electrospray tandem mass spectrometry. Phytochemistry 69:1773–1781

    Article  PubMed  CAS  Google Scholar 

  • Pavli OI, Panopoulos NJ, Goldbach R, Skaracis GN (2010) BNYVV-derived dsRNA confers resistance to rhizomania disease of sugar beet as evidenced by a novel transgenic hairy root approach. Transgenic Res

  • Peart JR, Cook G, Feys BJ, Parker JE, Baulcombe DC (2002) An EDS1 orthologue is required for N-mediated resistance against tobacco mosaic virus. Plant J 29:569–579

    Article  PubMed  CAS  Google Scholar 

  • Peltier C, Hleibieh K, Thiel H, Klein E, Bragard C, Gilmer D (2008) Molecular biology of the beet necrotic yellow vein virus. Plant Viruses 2:14–24

    Google Scholar 

  • Pfeffer S, Dunoyer P, Heim F, Richards KE, Jonard G, Ziegler-Graff V (2002) P0 of beet Western yellows virus is a suppressor of posttranscriptional gene silencing. J Virol 76:6815–6824

    Article  PubMed  CAS  Google Scholar 

  • Pollini CP, Masia A, Giunchedi L (1990) Free indole-3-acetic acid in sugar-beet root of rhizomania-susceptible and moderately resistant cultivars. Phytopathol Mediterr 29:191–195

    Google Scholar 

  • Prinsen E, Stijn Van L, Sevgi Ö, Henri Van O (2000) Auxin analysis. In: Iacobellis NS, Collmer A, Hutcheson SW, Mansfield JW, Morris CE, Murillo J, Schaad NW, Stead DE, Surico G, Ullrich M (eds) Plant hormone protocols. Kluwer Academic Humana Press, Dordrecht

    Google Scholar 

  • Quillet L, Guilley H, Jonard G, Richards K (1989) In vitro synthesis of biologically active beet necrotic yellow vein virus RNA. Virology 172:293–301

    Article  PubMed  CAS  Google Scholar 

  • Rahim MD, Andika IB, Han C, Kondo H, Tamada T (2007) RNA4-encoded p31 of beet necrotic yellow vein virus is involved in efficient vector transmission, symptom severity and silencing suppression in roots. J Gen Virol 88:1611–1619

    Article  PubMed  CAS  Google Scholar 

  • Schmidlin L, De Bruyne E, Weyens G, Lefebvre M, Gilmer D (2008) Identification of differentially expressed root genes upon rhizomania disease. Mol Plant Pathol 9:741–751

    Article  PubMed  CAS  Google Scholar 

  • Smets R, Claes V, Van Onckelen HA, Prinsen E (2003) Extraction and quantitative analysis of 1-aminocyclopropane-1-carboxylic acid in plant tissue by gas chromatography coupled to mass spectrometry. J Chromatogr A 993:79–87

    Article  PubMed  CAS  Google Scholar 

  • Staswick PE, Su W, Howell SH (1992) Methyl jasmonate inhibition of root growth and induction of a leaf protein are decreased in an Arabidopsis thaliana mutant. Proc Natl Acad Sci USA 89:6837–6840

    Article  PubMed  CAS  Google Scholar 

  • Tamada T, Abe H (1989) Evidence that beet necrotic yellow vein virus RNA-4 is essential for transmission by the fungus Polymyxa betae. J Gen Virol 70:3391–3398

    Article  CAS  Google Scholar 

  • Tamada T, Shirako Y, Abe H, Saito M, Kigushi T, Harada T (1989) Production and pathogenicity of isolates of Beet necrotic yellow vein virus with different numbers of RNA components. J Gen Virol 70:3399–3409

    Article  CAS  Google Scholar 

  • Tamada T, Uchino H, Kusume T, Saito M (1999) RNA 3 deletion mutants of beet necrotic yellow vein virus do not cause rhizomania disease in sugar beets. Phytopathology 89:1000–1006

    Article  PubMed  CAS  Google Scholar 

  • Thiel H, Varrelmann M (2009) Identification of beet necrotic yellow vein virus P25 pathogenicity factor-interacting sugar beet proteins that represent putative virus targets or components of plant resistance. Mol Plant Microbe Interact 22:999–1010

    Article  PubMed  CAS  Google Scholar 

  • Tsujimoto Y, Numaga T, Ohshima K, Yano MA, Ohsawa R, Goto DB, Naito S, Ishikawa M (2003) Arabidopsis tobamovirus multiplication (TOM) 2 locus encodes a transmembrane protein that interacts with TOM1. EMBO J 22:335–343

    Article  PubMed  CAS  Google Scholar 

  • Ulmasov T, Murfett J, Hagen G, Guilfoyle TJ (1997) Aux/IAA proteins repress expression of reporter genes containing natural and highly active synthetic auxin response elements. Plant Cell 9:1963–1971

    Article  PubMed  CAS  Google Scholar 

  • Van Der Straeten D, Djudzman A, Van Caeneghem W, Smalle J, Van Montagu M (1993) Genetic and physiological analysis of a new locus in Arabidopsis that confers resistance to 1-aminocyclopropane-1-carboxylic acid and ethylene and specifically affects the ethylene signal transduction pathway. Plant Physiol 102:401–408

    Google Scholar 

  • Vetter G, Hily JM, Klein E, Schmidlin L, Haas M, Merkle T, Gilmer D (2004) Nucleo-cytoplasmic shuttling of the beet necrotic yellow vein virus RNA-3-encoded p25 protein. J Gen Virol 85:2459–2469

    Article  PubMed  CAS  Google Scholar 

  • Voinnet O, Vain P, Angell S, Baulcombe DC (1998) Systemic spread of sequence-specific transgene RNA degradation in plants is initiated by localized introduction of ectopic promoterless DNA. Cell 95:177–187

    Article  PubMed  CAS  Google Scholar 

  • Voinnet O, Rivas S, Mestre P, Baulcombe D (2003) An enhanced transient expression system in plants based on suppression of gene silencing by the p19 protein of tomato bushy stunt virus. Plant J 33:949–956

    Article  PubMed  CAS  Google Scholar 

  • Yamanaka T, Ohta T, Takahashi M, Meshi T, Schmidt R, Dean C, Naito S, Ishikawa M (2000) TOM1, an Arabidopsis gene required for efficient multiplication of a tobamovirus, encodes a putative transmembrane protein. Proc Natl Acad Sci USA 97:10107–10112

    Article  PubMed  CAS  Google Scholar 

  • Yamanaka T, Imai T, Satoh R, Kawashima A, Takahashi M, Tomita K, Kubota K, Meshi T, Naito S, Ishikawa M (2002) Complete inhibition of tobamovirus multiplication by simultaneous mutations in two homologous host genes. J Virol 76:2491–2497

    Article  PubMed  CAS  Google Scholar 

  • Yoshida K, Kaothien P, Matsui T, Kawaoka A, Shinmyo A (2003) Molecular biology and application of plant peroxidase genes. Appl Microbiol Biotechnol 60:665–670

    PubMed  CAS  Google Scholar 

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Acknowledgments

The authors are grateful to Valérie Cognat for her help with statistical analysis of hormonal growth effects, Malek Alioua for qPCR advices and sequencing, Danièle Scheidecker for technical assistance and Thierry Heitz for helpful discussion on JA assays. EP performed hormonal assays of auxins, ACC and ABA. ME and DH performed TEM and JA assays, respectively. CP and LS were supported by SES-VanderHave France (Nérac) under CIFRE conventions.

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Authors and Affiliations

  1. Institut de biologie moléculaire des plantes, Laboratoire Propre du CNRS (UPR 2357) conventionné avec l’Université de Strasbourg, 12 rue du Général Zimmer, 67084, Strasbourg, France

    Claire Peltier, Laure Schmidlin, Elodie Klein, Mathieu Erhardt, Dimitri Heintz & David Gilmer

  2. Department of Biology, University of Antwerp, Groenenborgerlaan 171, 2020, Antwerpen, Belgium

    Els Prinsen

  3. SESVanderHave nv, Industriepark 15, Soldatenplein Z2, 3300, Tienen, Belgium

    Guy Weyens & Marc Lefebvre

  4. URGV, UMR INRA 1165, CNRS 8114, UEVE, 2 rue Gaston Crémieux, CP5708, 91057, Evry cedex, France

    Ludivine Taconnat & Jean-Pierre Renou

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  1. Claire Peltier
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  2. Laure Schmidlin
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Correspondence to David Gilmer.

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The authors Claire Peltier and Laure Schmidlin contributed equally to this work.

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11248_2010_9424_MOESM1_ESM.tif

Expression of the p25 protein allows germination of A. thaliana plants in non-permissive conditions. Seeds from wild-type Col-0 plants (WT) and p25 protein-expressing transgenic lines (35S::P25B, 35S::P25E and 35S::P25T) were sowed on control media (MS) or media containing MeJA (100 μM), SA (400 μM), ABA (300 nM) or ACC (50 μM). Seed germination and plant growth were compared after 14 days. (TIFF 5,753 kb)

11248_2010_9424_MOESM2_ESM.tif

Presentation of transcript profiling experiments comparing roots of wild-type A. thaliana plants (WT) and p25 protein-expressing transgenic 35S::P25B (B), 35S::P25E (E) and 35S::P25T (T) lines after 10 days (10d) or 17 days (17d) of sterile growth. (a) Picture and scheme of plant distribution on MS media followed to standardize growth conditions of wild-type and transgenic seedlings. The scheme does not correspond to the plate image presented but explain the alternative saw of both WT and transgenic seeds used. (b) Scheme of hybridization planning of the two-colour microarray analyses. (c) Circularization of the expression ratios (X + Z−Y−W) comparing wild-type plants (WT) and 35S::P25B plants (B) after 10 days (10d) and 17 days (17d) provides theoretically a zero value. Calculation with the entire set of 3,428 differentially deregulated genes provides a mean value equal to −0.03 with a standard deviation (SD) of 0.22. (TIFF 7,926 kb)

11248_2010_9424_MOESM3_ESM.tif

Functional GO biological process categorization of differentially regulated genes in roots of the transgenic 35S::P25B plants. Only genes with a differential expression equal to or higher than 0.61 or −0.61 were included in the analysis, resulting in 1,363 candidate genes (1,067 up-regulated and 296 down-regulated). (TIFF 6,223 kb)

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Peltier, C., Schmidlin, L., Klein, E. et al. Expression of the Beet necrotic yellow vein virus p25 protein induces hormonal changes and a root branching phenotype in Arabidopsis thaliana . Transgenic Res 20, 443–466 (2011). https://doi.org/10.1007/s11248-010-9424-3

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