Transcriptional and metabolic profiling of grape (Vitis vinifera L.) leaves unravel possible innate resistance against pathogenic fungi.

Grapevine species (Vitis sp.) are prone to several diseases, fungi being the major pathogens compromising its cultivation and economic profit around the world. Knowledge of the complexity of mechanisms responsible for resistance to fungus infection of cultivars, such as Regent, is necessary for strategies to be defined which will improve resistance in highly susceptible crop species. Transcript and metabolic profiles of the Vitis vinifera cultivars Regent and Trincadeira (resistant and susceptible to fungi, respectively) were analysed by cDNA microarray, quantitative real-time PCR, and nuclear magnetic resonance spectroscopy. The integration of datasets obtained through transcriptome and metabolome analysis revealed differences in transcripts and metabolites between both cultivars. These differences are probably associated with the innate resistance of Regent towards the mildews. Several transcripts related to stress and defence, namely a subtilisin-like protease, phenylalanine ammonia lyase, S-adenosylmethionine synthase, WD-repeat protein like, and J2P, were up-regulated in Regent suggesting an intrinsic resistance capability of this cultivar. A metabolic profile revealed an accumulation of compounds such as inositol and caffeic acid, which are known to confer resistance to fungi. The differences in transcripts and metabolites detected are discussed in terms of the metabolic pathways and their possible role in plant defence against pathogen attack, as well as their potential interest to discriminate among resistant and susceptible grapevine cultivars.

[1]  Jong-Bum Kim,et al.  EST analysis of genes involved in secondary metabolism in Camellia sinensis (tea), using suppression subtractive hybridization , 2004 .

[2]  T. Wilkins,et al.  A modified hot borate method significantly enhances the yield of high-quality RNA from cotton (Gossypium hirsutum L.). , 1994, Analytical biochemistry.

[3]  Robert Verpoorte,et al.  Metabolic Discrimination of Catharanthus roseus Leaves Infected by Phytoplasma Using 1H-NMR Spectroscopy and Multivariate Data Analysis1 , 2004, Plant Physiology.

[4]  C. Espinoza,et al.  Gene expression associated with compatible viral diseases in grapevine cultivars , 2007, Functional & Integrative Genomics.

[5]  Jason E. Stewart,et al.  Minimum information about a microarray experiment (MIAME)—toward standards for microarray data , 2001, Nature Genetics.

[6]  D. Merdinoglu,et al.  Four specific isogenes of the anthocyanin metabolic pathway are systematically co-expressed with the red colour of grape berries , 2006 .

[7]  A. Osbourn,et al.  Fungal Resistance to Plant Antibiotics as a Mechanism of Pathogenesis , 1999, Microbiology and Molecular Biology Reviews.

[8]  R. Henry,et al.  cDNA microarray analysis of developing grape (Vitis vinifera cv. Shiraz) berry skin , 2004, Functional & Integrative Genomics.

[9]  P. Vera,et al.  Identification of a New Pathogen-induced Member of the Subtilisin-like Processing Protease Family from Plants* , 1997, The Journal of Biological Chemistry.

[10]  T. Lithgow,et al.  The J‐protein family: modulating protein assembly, disassembly and translocation , 2004, EMBO reports.

[11]  K. Edwards,et al.  Quantitative trait locus analysis of fungal disease resistance factors on a molecular map of grapevine , 2004, Theoretical and Applied Genetics.

[12]  E. Maul,et al.  Genetic mapping and localization of quantitative trait loci affecting fungal disease resistance and leaf morphology in grapevine (Vitis vinifera L) , 2007, Molecular Breeding.

[13]  A. Wachter,et al.  Structural Basis for the Redox Control of Plant Glutamate Cysteine Ligase* , 2006, Journal of Biological Chemistry.

[14]  P. Vera,et al.  Primary structure and expression of a pathogen-induced protease (PR-P69) in tomato plants: Similarity of functional domains to subtilisin-like endoproteases. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[15]  G. Gibson,et al.  Microarray Analysis , 2020, Definitions.

[16]  Marianne Defernez,et al.  Factors affecting the robustness of metabolite fingerprinting using 1H NMR spectra. , 2003, Phytochemistry.

[17]  D. Galbraith,et al.  Methods for Transcriptional Profiling in Plants. Be Fruitful and Replicate , 2004, Plant Physiology.

[18]  Jean-Marc Nuzillard,et al.  NMR metabolomics to revisit the tobacco mosaic virus infection in Nicotiana tabacum leaves. , 2006, Journal of natural products.

[19]  J. Cushman,et al.  Water and salinity stress in grapevines: early and late changes in transcript and metabolite profiles , 2007, Functional & Integrative Genomics.

[20]  C. Ford,et al.  Consequences of Transferring Three Sorghum Genes for Secondary Metabolite (Cyanogenic Glucoside) Biosynthesis to Grapevine Hairy Roots , 2006, Transgenic Research.

[21]  Susan Flores,et al.  Overexpression of D-myo-inositol-3-phosphate synthase leads to elevated levels of inositol in Arabidopsis , 1997, Plant Molecular Biology.

[22]  D. Somers,et al.  Fungal development and induction of defense response genes during early infection of wheat spikes by Fusarium graminearum. , 2000, Molecular plant-microbe interactions : MPMI.

[23]  RAINER BREITLING,et al.  Rank-based Methods as a Non-parametric Alternative of the T-statistic for the Analysis of Biological Microarray Data , 2005, J. Bioinform. Comput. Biol..

[24]  P. Vera,et al.  Expression of subtilisin-like serine proteases in Arabidopsis thaliana is cell-specific and responds to jasmonic acid and heavy metals with developmental differences. , 2003, Physiologia plantarum.

[25]  R. Dixon.  Natural products and plant disease resistance , 2001, Nature.

[26]  S. Dudoit,et al.  Normalization for cDNA microarray data: a robust composite method addressing single and multiple slide systematic variation. , 2002, Nucleic acids research.

[27]  Daniel Kost,et al.  Low-frequency electromagnetic fields induce a stress effect upon higher plants, as evident by the universal stress signal, alanine. , 2003, Biochemical and biophysical research communications.

[28]  S. Delrot,et al.  Isogene specific oligo arrays reveal multifaceted changes in gene expression during grape berry (Vitis vinifera L.) development , 2004, Planta.

[29]  Chunzhao Liu,et al.  Light-enhanced caffeic acid derivatives biosynthesis in hairy root cultures of Echinacea purpurea , 2007, Plant Cell Reports.

[30]  R. Dixon,et al.  Phenylpropanoid compounds and disease resistance in transgenic tobacco with altered expression of L-phenylalanine ammonia-lyase. , 2003, Phytochemistry.

[31]  R. Dixon,et al.  Increased disease susceptibility of transgenic tobacco plants with suppressed levels of preformed phenylpropanoid products. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[32]  Y. Choi,et al.  Metabolic characterization of Brassica rapa leaves by NMR spectroscopy. , 2007, Journal of agricultural and food chemistry.

[33]  Rainer Breitling,et al.  The Potassium-Dependent Transcriptome of Arabidopsis Reveals a Prominent Role of Jasmonic Acid in Nutrient Signaling1[w] , 2004, Plant Physiology.

[34]  D B Kell,et al.  Detection of the dipicolinic acid biomarker in Bacillus spores using Curie-point pyrolysis mass spectrometry and Fourier transform infrared spectroscopy. , 2000, Analytical chemistry.

[35]  M. S. Pais,et al.  Cost effective method for construction of high quality cDNA libraries. , 2007, Biomolecular engineering.

[36]  G. Coruzzi,et al.  Use of Arabidopsis mutants and genes to study amide amino acid biosynthesis. , 1995, The Plant cell.

[37]  K. Herrmann.  The Shikimate Pathway: Early Steps in the Biosynthesis of Aromatic Compounds. , 1995, The Plant cell.

[38]  Rainer Breitling,et al.  Rank products: a simple, yet powerful, new method to detect differentially regulated genes in replicated microarray experiments , 2004, FEBS letters.

[39]  J. V. Van Beeumen,et al.  Molecular Phenotyping of the pal1 and pal2 Mutants of Arabidopsis thaliana Reveals Far-Reaching Consequences on Phenylpropanoid, Amino Acid, and Carbohydrate Metabolism , 2004, The Plant Cell Online.

[40]  K. Kohno,et al.  Antimicrobial activity of Polygonum tinctorium Lour: extract against oral pathogenic bacteria , 2006, Journal of Natural Medicines.

[41]  A. Comeau,et al.  Metabolic profiling and factor analysis to discriminate quantitative resistance in wheat cultivars against fusarium head blight , 2005 .

[42]  John M Baker,et al.  Recent applications of NMR spectroscopy in plant metabolomics , 2007, The FEBS journal.

[43]  D. Rhodes,et al.  Effects of heat shock on amino Acid metabolism of cowpea cells. , 1990, Plant physiology.

[44]  S. Roje.  S-Adenosyl-L-methionine: beyond the universal methyl group donor. , 2006, Phytochemistry.

[45]  王毅,et al.  棉花（Gossypium hirsutum L.）花粉壁的发育 , 1994 .

[46]  Jonathan D. G. Jones,et al.  The plant proteolytic machinery and its role in defence. , 2004, Current opinion in plant biology.

[47]  C. Nautiyal,et al.  Induction of Plant Defense Enzymes and Phenolics by Treatment With Plant Growth–Promoting Rhizobacteria Serratia marcescens NBRI1213 , 2006, Current Microbiology.

[48]  Kazuo Shinozaki,et al.  Stable isotope labeling of Arabidopsis thaliana for an NMR-based metabolomics approach. , 2004, Plant & cell physiology.

[49]  N. Kruger,et al.  Metabolite fingerprinting and profiling in plants using NMR. , 2004, Journal of experimental botany.

[50]  A. Smigocki,et al.  Insect feeding-induced differential expression of Beta vulgaris root genes and their regulation by defense-associated signals , 2006, Plant Cell Reports.

[51]  T. Wolpert,et al.  Purification and Characterization of Serine Proteases That Exhibit Caspase-Like Activity and Are Associated with Programmed Cell Death in Avena sativa , 2004, The Plant Cell Online.

[52]  T. Widmer,et al.  Plant extracts containing caffeic acid and rosmarinic acid inhibit zoospore germination of Phytophthora spp. pathogenic to Theobroma cacao , 2006, European Journal of Plant Pathology.

[53]  S. Roje.  S-Adenosyl-L-methionine: Beyond the Universal Methyl Group Donor , 2006 .

[54]  R. Eibach,et al.  SUCCESS IN RESISTANCE BREEDING: "REGENT" AND ITS STEPS INTO THE MARKET , 2003 .

[55]  N. Medina-Escobar,et al.  Cloning, expression and immunolocalization pattern of a cinnamyl alcohol dehydrogenase gene from strawberry (Fragaria x ananassa cv. Chandler). , 2002, Journal of experimental botany.

[56]  P. Vera,et al.  A Genomic Cluster Containing Four Differentially Regulated Subtilisin-like Processing Protease Genes Is in Tomato Plants* , 1999, The Journal of Biological Chemistry.

[57]  A. Kortekamp,et al.  Characterization of Plasmopara-resistance in grapevine using in vitro plants. , 2003, Journal of plant physiology.