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Fast induction of biosynthetic polysaccharide genes lpxA, lpxE, and rkpI of Rhizobium sp. strain PRF 81 by common bean seed exudates is indicative of a key role in symbiosis

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Abstract

Rhizobial surface polysaccharides (SPS) are, together with nodulation (Nod) factors, recognized as key molecules for establishment of rhizobia–legume symbiosis. In Rhizobium tropici, an important nitrogen-fixing symbiont of common bean (Phaseolus vulgaris L.), molecular structures and symbiotic roles of the SPS are poorly understood. In this study, Rhizobium sp. strain PRF 81 genes, belonging to the R. tropici group, were investigated: lpxA and lpxE, involved in biosynthesis and modification of the lipid-A anchor of lipopolysaccharide (LPS), and rkpI, involved in synthesis of a lipid carrier required for production of capsular polysaccharides (KPS). Reverse transcription quantitative PCR (RT-qPCR) analysis revealed, for the first time, that inducers released from common bean seeds strongly stimulated expression of all three SPS genes. When PRF 81 cells were grown for 48 h in the presence of seed exudates, twofold increases (p < 0.05) in the transcription levels of lpxE, lpxA, and rkpI genes were observed. However, higher increases (p < 0.05) in transcription rates, about 50-fold for lpxE and about 30-fold for lpxA and rkpI, were observed after only 5 min of incubation with common bean seed exudates. Evolutionary analyses revealed that lpxA and lpxE of PRF81 and of the type strain of R. tropici CIAT899Tclustered with orthologous Rhizobium radiobacter and were more related to R. etli and Rhizobium leguminosarum, while rkpI was closer to the Sinorhizobium sp. group. Upregulation of lpxE, lpxA, and rkpI genes suggests that seed exudates can modulate production of SPS of Rhizobium sp. PRF81, leading to cell wall changes necessary for symbiosis establishment.

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Acknowledgments

The study was partially supported by the CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico, Brazil) (Repensa, 557746/2009-4). L. R. Oliveira acknowledges an M.Sc. fellowship (552874/2008-6) and M. Hungria, a research fellowship (300547/2010-2) from CNPq. The authors thank Allan R.J. Eaglesham for help with the manuscript. Manuscript approved for publication by the Editorial Board of Embrapa Soja as manuscript 18/2012.

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Authors and Affiliations

  1. Embrapa Soja, Cx Postal 231, 86001-970, Londrina, Paraná, Brazil

    Luciana Ruano Oliveira, Francismar Corrêa Marcelino-Guimarães & Mariangela Hungria

  2. Department of Microbiology, Universidade Estadual de Londrina, Londrina, Brazil

    Luciana Ruano Oliveira & Mariangela Hungria

  3. Department of Biology, Universidade Estadual de Londrina, Londrina, Brazil

    Elisete Pains Rodrigues

  4. Department of Biochemistry and Biotechnology, Universidade Estadual de Londrina, Londrina, Brazil

    André Luiz Martinez Oliveira

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  1. Luciana Ruano Oliveira
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Correspondence to Mariangela Hungria.

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Luciana Ruano Oliveira and Elisete Pains Rodrigues contributed equally to this work.

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Supplementary fig. 1

Unrooted phylogenetic analysis (top) and genomic arrangement (bottom) of lpxE gene from closely related bacteria. Neighbor-joining analysis was based on ClustalW alignment of translated nucleotide sequences (accession numbers are provided as supplementary file) using the maximum composite likelihood method. The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (2,000 replicates) is shown above the branches. Only bootstrapping values >80 are shown. Evolutionary analyses were performed in MEGA 5 (Tamura et al. 2011; Saitou and Nei 1987; Tamura et al. 2004; Felsenstein 1985) (DOCX 145 kb)

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Oliveira, L.R., Rodrigues, E.P., Marcelino-Guimarães, F.C. et al. Fast induction of biosynthetic polysaccharide genes lpxA, lpxE, and rkpI of Rhizobium sp. strain PRF 81 by common bean seed exudates is indicative of a key role in symbiosis. Funct Integr Genomics 13, 275–283 (2013). https://doi.org/10.1007/s10142-013-0322-7

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