Functional Genomics Plant-Pathogen

Summary

Research projects in our laboratory are broadly focused on different aspects of the molecular mechanisms of defence and pathogenesis of diseases induced by virus and bacteria in plants.

Research Lines

Functional genomics applied to the study of PVX and Xanthomonas pathogenesis in higher plants

1- Functional genomics applied to the study of PVX pathogenesis in Solanum tuberosum The goal of this project is to understand the biochemistry and genetic bases of resistance to potato virus X (PVX) mediated by the resistance gene Nb in S. tuberosum. Chromosome V of wild and cultivated potato contains a cluster of R-genes encoding resistance to diverse pathogens. The short arm of this chromosome harbors genes encoding resistance to Phytophtora infestans (R1) and hypersensitive resistance to the ROTH 1 strain of Potato virus X (Nb). Our research group as part of a joint effort with the NSF Potato Genome Project has constructed an integrated geneticphysical map of the Nb-R1 region using molecular markers from the tetraploid cultivar Pentland Ivory (genotype Nb, R1) and the allohexaploid wild species Solanum demissum. Sequence analysis of the region (~ 2 Mb) corresponding to the three haplotypes of Solanum demissum predicts the presence of more than 40 RGCs (resistance gene candidates) belonging to three distinct resistance-gene families (potato R1; tomato Prf and Bs4). Genetic and molecular analysis show that one of the RGCs sequences belongs to Nb. Our laboratory had also developed an agroinfiltration system to assess the transient expression of the viral elicitor, the 25 kDa protein, which will be used for functional complementation analysis of Nb-RGC. Towards the goal of elucidating defense signaling pathways, we have also studied the role of salicylic acid (SA) in Nb-mediated resistance to PVX infection, we have shown that SA is involved in the defense response mediated by Nb in Solanum tuberosum but the role played is notably different to that reported for the experimental model Arabidopsis thaliana and Nicotiana tabacum. 2- Functional genomics applied to the study of Xanthomonas pathogenesis in Citrus limon Xac is responsible for the canker disease affecting citrus plants throughout the world. The goal of this study is to generate knowledge for the development of crop protection methods based on interference with key biological and molecular processes in the epiphytic and disease phases of bacterial pathogenesis. We have studied the formation of biofilms of Xac and its role in the epiphytic survival and canker pathogenicity. We have shown that Xac develop biofilm on either abiotic or biotic surfaces and the presence of plant-associated biofilms is correlated with the bacterial survival and Xac pathogenicity. We have shown by genetic, biochemistry and molecular analysis that the biofilm formation of Xac requires xanthan production. Without xanthan, the bacteria are unable to retain water and cannot withstand abiotic stress, and thus cannot survive on the leaf surface. Based in our results we have established a Xac - Citrus limon assay and have used it to identify differential gene expression during Xac - Citrus interaction. In addition, we are developing genomic tools to study the function of these isolated genes in citrus.

Selected Publications

  • Sendín L., Filippone M.P., Orce I.G., Enrique R., Rigano L., Peña L., Vojnov A.A., Marano M.R. and Castagnaro A.P. (2011) Transient expression of Capsicum chacoense Bs2 gene in Citrus limon as an approach for management of citrus canker disease. Plant Pathology (en prensa).
  • Malamud F., Torres P.S., Roeschlin R., Rigano L.A., Enrique R., Bonomi H.R., Castagnaro A.P., Marano M.R., Vojnov A.A. (2011) Xanthomonas axonopodis pv. citri flagellum is required for mature biofilm and canker development. Microbiology, 157, 819-829. DOI 10.1099/mic.0.044255-0.
  • Enrique R., Siciliano F., Favaro M.A., Gerhardt N., Roeschlin R., Rigano L., Sendin L., Castagnaro A., Vojnov A. and Marano M.R. (2011) Novel demonstration of RNAi in citrus reveals importance of citrus callose synthase in defence against Xanthomonas citri subsp. citri. Plant Biotech. J. 9, 394-407.
  • Rigano L., Marano M.R., Castagnaro A.P., Do Amaral A.M. and Vojnov A.A. (2010) Rapid and sensitive detection of Citrus Bacterial Canker by loop-mediated isothermal amplification combined with simple visual evaluation methods. BMC Microbiology 10, 176. http://www.biomedcentral.com/1471-2180/10/176.
  • Vojnov A.A., do Amaral A.M., Dow J.M., Castagnaro A.P., and Marano M.R. (2010) Bacteria causing important diseases of citrus utilize distinct modes of pathogenesis to attack a common host. Appl Microbiol Biotechnol, DOI 10.1007/s00253-010-2631-2.
  • Sánchez G., Gerhardt N., Siciliano F., Vojnov A.A., Malcuit I. and Marano M.R. (2010) Salicylic acid is involved in the Nb-mediated defense responses to PVX in Solanum tuberosum. Mol. Plant-Microbe Interact. 23, 394-405.

Collaborators

  • Dr. Adrían A. Vojnov. Fundación Pablo Cassará, Centro de Ciencia y Tecnología “Dr. Cesar Milstein”, Saladillo 2468 C1440FFX, Ciudad de Buenos Aires, Argentina.
  • Dr. Atilio P. Castagnaro. Estación Experimental Agroindustrial Obispo Colombres, Casilla No. 9, Las Talitas, 4101, Tucumán, Argentina.
  • Dr. Norberto Gariglio. Facultad de Ciencias Agrarias, Universidad Nacional del Litoral, Esperanza, Santa Fe.
  • Dr. Vicente Conejero. Instituto de Biología Molecular y Celular de Plantas, Universidad Politécnica de Valencia, Valencia, España.
  • Dr. José Gadea Vacas. Instituto de Biología Molecular y Celular de Plantas, Universidad Politécnica de Valencia, Valencia, España.
  • Dr. Frederick G. Gmitter Jr. Centro de Educación e Investigación de los cítricos (CREC), Universidad de Florida, Estados Unidos.
  • Dr. J Maxwell Dow. BIOMERIT Research Centre, Department of Microbiology, National University of Ireland, Cork, Ireland.
  • Dra. Isabelle Malcuit. Algentech Ltd, Institute of Food Research. IFR2 Norwich Research Park, Norwich NR4 7UA, United Kingdom.

Grants

  • International Citrus Genome Consortium (ICGC): Providing tools to address HLB and other challenges. Florida Citrus Advanced Technology Programm - Control of Citrus Greening, Canker and Emerging Diseases of Citrus.
  • Programa de Cooperación Científico-Tecnológica entre MINCYT y MICINN (España) ES/09/08. “Genómica Funcional en Cítricos: Análisis de la respuesta frente a patógenos de gran incidencia en la citricultura de Argentina y España”.
  • Agencia Nacional de Promoción Científica y Tecnológica (ANPCyT).

Director

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Marano, María Rosa
Core CCT
Email: marano@ibr-conicet.gov.ar
Phone: +54 341 4237070
Office Extension: 653
Laboratory Extension: 624

Postdoctoral fellows

  • Lucila Garcia

Doctoral fellows

  • Sabrina E. Taselli
  • Facundo Uviedo
  • Ma. Celeste Molina

Imágenes

Silenciamiento génico mediante expresión transitoria del ARN doble cadena, inductor de ARNi. (A) Fenotipo de plantas de Citrus limón silenciadas en la fitoenodesaturasa (PDS) a 60 y 70 días post inoculación. (B) Magnificación de las hojas de (A), mostrando los diferentes fenotipos de silenciamiento en PDS.

La expresión transitoria de la proteína de 25-kDa de PVX induce la HR mediada por Nb en S. tuberosum (A), Fenotipo HR, hojas de papa cv Pentland Ivory NahG (Nb), no transformada (Nt, Nb) y susceptible (nb) infiltradas con Agrobacterium tumefanciens (transformado con el plásmido pBIN25K1) (B), Tinción con DAB o (C), azul de anilina para detectar la acumulación de H2O2 (precipitado marrón) y la deposición de calosa (puntos blancos), respectivamente, a 16 hpi.