of Sinorhizobium meliloti genes relevant for the colonization of host plant roots during the early stages of the symbiotic interaction

LOZANO MAURICIO JAVIER; SALAS MARÍA EUGENIA; PARISI, GUSTAVO; LOPEZ, JOSE LUIS; ALBICORO, FRANCISCO; MARTINI MARÍA CARLA; SALTO ILEANA PAULA; PISTORIO MARIANO; DEL PAPA MARÍA FLORENCIA; BECKER, ANKE; LAGARES ANTONIO

Rodas

Congreso; XVI International Congress on Molecular Plant - Microbe Interactions.; 2014

International Society for Molecular Plant-Microbe Interactions

375 IDENTIFICATION OF SINORHIZOBIUM MELILOTI GENES RELEVANT FOR THE COLONIZATION OF HOST PLANT ROOTS DURING THE EARLY STAGES OF THE SYMBIOTIC INTERACTION Mauricio Javier Lozano , Maria Eugenia Salas , Gustavo Parisi , José Luis Lopez , Francisco Javier Albicoro , Maria Carla Martini , Ileana Paula Salto , Mariano Pistorio , María Florencia Del Papa , Anke Becker , Antonio Lagares IBBM-Instituto de Biotecnología y Biología Molecular, Facultad de Ciencias Exactas, Universidad Nacional de La Plata, La Plata, Argentina bUniversidad Nacional de Quilmes, Centro de Estudios e Investigaciones, Bernal, Argentina cSYNMIKRO-Zentrum für Synthetische Mikrobiologie, Philipps Universität, Marburg, Germany Sinorhizobium meliloti is a gram-negative proteobacterium able to establish nitrogen-fixing symbiosis with legumes of the genera Medicago, Melilotus, and Trigonella. This association is tightly regulated, and rhizobial strains in the soil compete for the colonization of the symbiotic niche. Unfortunately, the molecular bases of the early competition are not fully understood yet, limiting the biotechnological improvement of inoculant strains. To discover new genes relevant for these early events we used two experimental approaches designed for the study of biological niches which cannot be easily replicated in vitro. Namely, Recombination-based In Vivo Expression Technology (RIVET) and Signature-Tagged Mutagenesis (STM). For the RIVET approach we generated a S. meliloti library of transcriptional fusions to tnpR (site-specific recombinase) and upon transferring it to the positive selection strain S. meliloti 2011R1ΩNGG (Lozano et al. 2011), we conducted several root colonization experiments using low numbers of bacteria (105 /ml - 106 /ml) in order to identify transcriptional fusions induced during the experiment. For the STM approach we used a large collection of Tn5 tagged mutants (Pobigaylo et al. 2006) using the same experimental design. To determine and compare the proportion mutants between the input (inoculum) and the output (root associated rhizobia) conditions we used microarrays and Illumina HiSeq sequencing platform. Both approaches were able to identify genes (pyrE, metA, SM2011_c04258) that might be of relevance for the early root colonization. Of the identified genes, only few (pyrE, metA) have been previously described as involved in nodule competition (Pobigaylo et al.). Lozano, M. J., M. E. Salas, M. A. Giusti, W. O. Draghi, G. A. Torres Tejerizo, M. C. Martini, M. F. Del Papa, M. Pistorio y A. Lagares (2011). "Development of new positive-selection RIVET tools: detection of induced promoters by the excision-based transcriptional activation of an aacCI (GmR)-gfp fusion." J Biotechnol 155(2): 147-155. Pobigaylo, N., S. Szymczak, T. W. Nattkemper y A. Becker (2008). "Identification of genes relevant to symbiosis and competitiveness in Sinorhizobium meliloti using signature-tagged mutants." Mol Plant Microbe Interact 21(2): 219-231. Pobigaylo, N., D. Wetter, S. Szymczak, U. Schiller, S. Kurtz, F. Meyer, T. W. Nattkemper y A. Becker (2006). "Construction of a large signature-tagged mini-Tn5 transposon library and its application to mutagenesis of Sinorhizobium meliloti." Appl Environ Microbiol 72(6): 4329-4337.