High-Resolution Transcriptomic Analyses of Sinorhizobium sp. NGR234 Bacteroids in Determinate Nodules of Vigna unguiculata and Indeterminate Nodules of Leucaena leucocephala

Abstract

The rhizobium-legume symbiosis is a model system for studying mutualistic interactions between bacteria and eukaryotes. Sinorhizobium sp. NGR234 is distinguished by its ability to form either indeterminate nodules or determinate nodules with diverse legumes. Here, we presented a high-resolution RNA-seq transcriptomic analysis of NGR234 bacteroids in indeterminate nodules of Leucaena leucocephala and determinate nodules of Vigna unguiculata. In contrast to exponentially growing free-living bacteria, non-growing bacteroids from both legumes recruited several common cellular functions such as cbb3 oxidase, thiamine biosynthesis, nitrate reduction pathway (NO-producing), succinate metabolism, PHB (poly-3-hydroxybutyrate) biosynthesis and phosphate/phosphonate transporters. However, different transcription profiles between bacteroids from two legumes were also uncovered for genes involved in the biosynthesis of exopolysaccharides, lipopolysaccharides, T3SS (type three secretion system) and effector proteins, cytochrome bd ubiquinol oxidase, PQQ (pyrroloquinoline quinone), cytochrome c550, pseudoazurin, biotin, phasins and glycolate oxidase, and in the metabolism of glutamate and phenylalanine. Noteworthy were the distinct expression patterns of genes encoding phasins, which are thought to be involved in regulating the surface/volume ratio of PHB granules. These patterns are in good agreement with the observed granule size difference between bacteroids from L. leucocephala and V. unguiculata.

RNA Sequencing Analysis of the Broad-Host-Range Strain Sinorhizobium fredii NGR234 Identifies a Large Set of Genes Linked to Quorum Sensing-Dependent Regulation in the Background of a traI and ngrI Deletion Mutant

“…Our data are in agreement with earlier findings reported for planktonic Bradyrhizobium japonicum cells and for S. meliloti suggesting that flagella play a pivotal role during root colonization but that they are most likely not essential during the infection process (56)(57)(58)(59). This hypothesis fits well with the recent observation that flagellar genes are downregulated in bacteroids in NGR234 (29) and with the observation that flagellar genes are subject to QS-mediated regulation in S. meliloti (60,61).…”

“…Recently, it was also shown that the T3SS-I locus was upregulated in the Leucaena leucocephala and V. unguiculata bacteroids. Transcription was, however, highest in the L. leucocephala nodules in comparison to the V. unguiculata nodules (29). Within this framework, our transcriptome study suggests that the T3SS-I is subject to a QS-independent regulation.…”

“…In contrast to the T3SS-I, not much is known about the function of the second T3SS locus. Only a few of the T3SS-II-associated genes were upregulated in L. leucocephala bacteroids (29), and a deletion of NGR_b22890 to NGR_b22950 in the T3SS-II did not result in a symbiotic phenotype, suggesting that the T3SS-II locus is not required for effective nodulation of the few legumes tested (13,88). In contrast to the QS-independent regulation of T3SS-I, the T3SS-II, however, appears to be regulated in a QS-dependent manner.…”

“…Interestingly, no study has yet focused on the QS-dependent gene regulation in sinorhizobia or closely related species using RNA-seq. Only very recently, an RNA-seq analysis was published on NGR234 with respect to a genome-wide change of the expression profile of the symbiotic lifestyle in bacteroids of two legume plants (29). This study has already given us a very detailed insight into the complexity of many processes linked to the bacterial infection process.…”

Genetic Analysis Reveals the Essential Role of Nitrogen Phosphotransferase System Components in Sinorhizobium fredii CCBAU 45436 Symbioses with Soybean and Pigeonpea Plants

“…Plant shoots were dried at 65°C for 5 days and then used for determining shoot dry weight per plant. For electron microscopy, ultrathin sections of 1-month nodules from soybeans were prepared and observed in a JEM-1230 transmission electron microscope as described earlier (31). To check nodule occupation, S. fredii mutants were individually mixed with the wild-type strain (or the mutant indicated) in an equal quantity (OD 600 , 0.2; 1-ml volume) and inoculated on soybeans.…”