Abstract
The legume symbiont Sinorhizobium meliloti expresses a plethora of small noncoding RNAs (sRNAs) whose function is mostly unknown. Here, we have functionally characterized two tandemly encoded S. meliloti Rm1021 sRNAs that are similar in sequence and structure. Homologous sRNAs (designated AbcR1 and AbcR2) have been shown to regulate several ABC transporters in the related α-proteobacteria Agrobacterium tumefaciens and Brucella abortus. In Rm1021, AbcR1 and AbcR2 exhibit divergent unlinked regulation and are stabilized by the RNA chaperone Hfq. AbcR1 is transcribed in actively dividing bacteria, either in culture, rhizosphere or within the invasion zone of mature alfalfa nodules. Conversely, AbcR2 expression is induced upon entry into stationary phase and under abiotic stress. Only deletion of AbcR1 resulted into a discrete growth delay in rich medium, but both are dispensable for symbiosis. Periplasmic proteome profiling revealed down-regulation of the branched-chain amino acid binding protein LivK by AbcR1, but not by AbcR2. A double-plasmid reporter assay confirmed the predicted specific targeting of the 5′-untranslated region of the livK mRNA by AbcR1 in vivo. Our findings provide evidences of independent regulatory functions of these sRNAs, probably to fine-tune nutrient uptake in free-living and undifferentiated symbiotic rhizobia.
Highlights
Sinorhizobium meliloti is a representative of the group of soildwelling a-proteobacteria, collectively known as rhizobia, which have the ability to engage in species-specific nitrogen-fixing endosymbioses with leguminous plants
Western-blot probing of protein extracts from the whole set of S. meliloti Rm1021 reporter strains grown to log phase in TY broth to detect the LivK::EGFP fusion protein fully confirmed the colony fluorescence observations (Fig. 6D). These results suggest that the AbcR1-mediated control of LivK is exerted by a canonical antisense interaction between the small noncoding RNAs (sRNAs) and the 59-untranslated region (UTR) of the mRNA, which probably results in occlusion of the ribosome binding site (RBS) and translational inhibition
We have addressed the functional characterization of the S. meliloti homologous trans-acting AbcR1 and AbcR2 sRNAs
Summary
Sinorhizobium meliloti is a representative of the group of soildwelling a-proteobacteria, collectively known as rhizobia, which have the ability to engage in species-specific nitrogen-fixing endosymbioses with leguminous plants. The outcome of these mutualistic plant-microbe interactions is the formation of root nodule structures in the host. Hfq binds diverse RNA molecules, including regulatory small noncoding RNAs (sRNAs) It has emerged as a global post-transcriptional regulator of gene expression with a great impact on bacterial physiology [14]
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