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Sinorhizobium meliloti DnaJ Is Required for Surface Motility, Stress Tolerance, and for Efficient Nodulation and Symbiotic Nitrogen Fixation.

ABSTRACT: Bacterial surface motility is a complex microbial trait that contributes to host colonization. However, the knowledge about regulatory mechanisms that control surface translocation in rhizobia and their role in the establishment of symbiosis with legumes is still limited. Recently, 2-tridecanone (2-TDC) was identified as an infochemical in bacteria that hampers microbial colonization of plants. In the alfalfa symbiont Sinorhizobium meliloti, 2-TDC promotes a mode of surface motility that is mostly independent of flagella. To understand the mechanism of action of 2-TDC in S. meliloti and unveil genes putatively involved in plant colonization, Tn5 transposants derived from a flagellaless strain that were impaired in 2-TDC-induced surface spreading were isolated and genetically characterized. In one of the mutants, the gene coding for the chaperone DnaJ was inactivated. Characterization of this transposant and newly obtained flagella-minus and flagella-plus dnaJ deletion mutants revealed that DnaJ is essential for surface translocation, while it plays a minor role in swimming motility. DnaJ loss-of-function reduces salt and oxidative stress tolerance in S. meliloti and hinders the establishment of efficient symbiosis by affecting nodule formation efficiency, cellular infection, and nitrogen fixation. Intriguingly, the lack of DnaJ causes more severe defects in a flagellaless background. This work highlights the role of DnaJ in the free-living and symbiotic lifestyles of S. meliloti.

SUBMITTER: Brito-Santana P

PROVIDER: S-EPMC10055731 | biostudies-literature | 2023 Mar

REPOSITORIES: biostudies-literature

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<i>Sinorhizobium meliloti</i> DnaJ Is Required for Surface Motility, Stress Tolerance, and for Efficient Nodulation and Symbiotic Nitrogen Fixation.

Brito-Santana Paula P Duque-Pedraza Julián J JJ Bernabéu-Roda Lydia M LM Carvia-Hermoso Cristina C Cuéllar Virginia V Fuentes-Romero Francisco F Acosta-Jurado Sebastián S Vinardell José-María JM Soto María J MJ

International journal of molecular sciences 20230319 6

Bacterial surface motility is a complex microbial trait that contributes to host colonization. However, the knowledge about regulatory mechanisms that control surface translocation in rhizobia and their role in the establishment of symbiosis with legumes is still limited. Recently, 2-tridecanone (2-TDC) was identified as an infochemical in bacteria that hampers microbial colonization of plants. In the alfalfa symbiont <i>Sinorhizobium meliloti</i>, 2-TDC promotes a mode of surface motility that ...[more]

PMID: 36982921

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Project description:Symbiotic nitrogen fixation (SNF) is an energetically expensive process performed by bacteria during endosymbiotic relationships with plants. The bacteria require the plant to provide a carbon source for the generation of reductant to power SNF. While C4-dicarboxylates (succinate, fumarate, and malate) appear to be the primary, if not sole, carbon source provided to the bacteria, the contribution of each C4-dicarboxylate is not known. We address this issue using genetic and systems-level analyses. Expression of a malate-specific transporter (MaeP) in Sinorhizobium meliloti Rm1021 dct mutants unable to transport C4-dicarboxylates resulted in malate import rates of up to 30% that of the wild type. This was sufficient to support SNF with Medicago sativa, with acetylene reduction rates of up to 50% those of plants inoculated with wild-type S. melilotiRhizobium leguminosarum bv. viciae 3841 dct mutants unable to transport C4-dicarboxylates but expressing the maeP transporter had strong symbiotic properties, with Pisum sativum plants inoculated with these strains appearing similar to plants inoculated with wild-type R. leguminosarum This was despite malate transport rates by the mutant bacteroids being 10% those of the wild type. An RNA-sequencing analysis of the combined P. sativum-R. leguminosarum nodule transcriptome was performed to identify systems-level adaptations in response to the inability of the bacteria to import succinate or fumarate. Few transcriptional changes, with no obvious pattern, were detected. Overall, these data illustrated that succinate and fumarate are not essential for SNF and that, at least in specific symbioses, l-malate is likely the primary C4-dicarboxylate provided to the bacterium.IMPORTANCE Symbiotic nitrogen fixation (SNF) is an economically and ecologically important biological process that allows plants to grow in nitrogen-poor soils without the need to apply nitrogen-based fertilizers. Much research has been dedicated to this topic to understand this process and to eventually manipulate it for agricultural gains. The work presented in this article provides new insights into the metabolic integration of the plant and bacterial partners. It is shown that malate is the only carbon source that needs to be available to the bacterium to support SNF and that, at least in some symbioses, malate, and not other C4-dicarboxylates, is likely the primary carbon provided to the bacterium. This work extends our knowledge of the minimal metabolic capabilities the bacterium requires to successfully perform SNF and may be useful in further studies aiming to optimize this process through synthetic biology approaches. The work describes an engineering approach to investigate a metabolic process that occurs between a eukaryotic host and its prokaryotic endosymbiont.

Project description:BACKGROUND: Sinorhizobium meliloti is a model system for the studies of symbiotic nitrogen fixation. An extensive polymorphism at the genetic and phenotypic level is present in natural populations of this species, especially in relation with symbiotic promotion of plant growth. AK83 and BL225C are two nodule-isolated strains with diverse symbiotic phenotypes; BL225C is more efficient in promoting growth of the Medicago sativa plants than strain AK83. In order to investigate the genetic determinants of the phenotypic diversification of S. meliloti strains AK83 and BL225C, we sequenced the complete genomes for these two strains. RESULTS: With sizes of 7.14 Mbp and 6.97 Mbp, respectively, the genomes of AK83 and BL225C are larger than the laboratory strain Rm1021. The core genome of Rm1021, AK83, BL225C strains included 5124 orthologous groups, while the accessory genome was composed by 2700 orthologous groups. While Rm1021 and BL225C have only three replicons (Chromosome, pSymA and pSymB), AK83 has also two plasmids, 260 and 70 Kbp long. We found 65 interesting orthologous groups of genes that were present only in the accessory genome, consequently responsible for phenotypic diversity and putatively involved in plant-bacterium interaction. Notably, the symbiosis inefficient AK83 lacked several genes required for microaerophilic growth inside nodules, while several genes for accessory functions related to competition, plant invasion and bacteroid tropism were identified only in AK83 and BL225C strains. Presence and extent of polymorphism in regulons of transcription factors involved in symbiotic interaction were also analyzed. Our results indicate that regulons are flexible, with a large number of accessory genes, suggesting that regulons polymorphism could also be a key determinant in the variability of symbiotic performances among the analyzed strains. CONCLUSIONS: In conclusions, the extended comparative genomics approach revealed a variable subset of genes and regulons that may contribute to the symbiotic diversity.

Project description:Differences in genome size and gene content are among the most important signatures of microbial adaptation and genome evolution. Here, we investigated the patterns of genome variation among strains of the symbiotic nitrogen-fixing bacterium Sinorhizobium meliloti. Using the sequenced strain Rm1021 as a reference, the genome size and gene content variations were analyzed among ten diverse natural strains, through pulse field gel electrophoresis (PFGE) and whole-genome microarray hybridizations. Our PFGE analysis showed a genome size range of 6.45-7.01Mbp, with the greatest variation arising from the pSymA replicon, followed by that of pSymB. No observable size difference was evident among the chromosomes. Consistent with this pattern of size differences, 41.2% of ORFs on pSymA were variably absent/present, followed by 12.7% on pSymB, and 3.7% on the chromosome. However, the percentages of ORFs that were variably duplicated were more evenly distributed among the three replicons, 11.0%, 16.5% and 15.3% respectively for ORFs on pSymA, pSymB and the chromosome. Among the 10 strains, the percentages of absent ORFs ranged from 1.51% to 6.35% and those of duplicated ORFs ranged from 0.27% to 8.56%. Our analyses showed that host plants, geographic origins, multilocus enzyme electrophoretic types, and replicon sizes had little influence on the distribution patterns of absent or duplicated ORFs. The proportions of ORFs that were either variably absent/present or variably duplicated differed greatly among the functional categories, for each of the three replicons as well as for the whole genome. Interestingly, we observed positive correlations among the three replicons in their numbers of absent ORFs as well as the numbers of duplicated ORFs, consistent with coordinated gene gains/losses in this important bacterium in nature. microarray:Sm6kOligo A total of 12 strains were included for the microarray analyses in this study. The 12 strains included two reference strains and ten other natural strains. Reference strain Rm1021 was the strain with a completely sequenced genome, from which the whole genome microarray was based on (Krol et al. 2004). The second reference strain RmF909 was derived from strain Rm1021 but with a deletion of 575 ORFs located on the megaplasmid pSymB (genomic location 106724-735431, Charles et al. 1991). The two reference strains were used for positive and negative controls in our microarray hybridization experiments and for establishing threshold values for determining whether an ORF is present or absent among natural strains.

Dataset Information

Sinorhizobium meliloti DnaJ Is Required for Surface Motility, Stress Tolerance, and for Efficient Nodulation and Symbiotic Nitrogen Fixation.

Publications

<i>Sinorhizobium meliloti</i> DnaJ Is Required for Surface Motility, Stress Tolerance, and for Efficient Nodulation and Symbiotic Nitrogen Fixation.

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