RNase III influences microaerobic symbiotic pathways and RNA regulation in Sinorhizobium meliloti

Abstract

Bacterial ribonucleases (RNases) are central components of post-transcriptional networks underlying environmental adaptation. However, their contribution to the ecological specialization of bacteria with complex lifestyles, such as nitrogen-fixing legume symbionts, remains poorly understood. Here, we investigated the role of the double-stranded RNase III ortholog (SmRNase III) in Sinorhizobium meliloti, the symbiotic partner of alfalfa (Medicago sativa L.). Loss of SmRNase III function affected the expression of nearly 30% of protein-coding genes and 12% of annotated non-coding RNAs (sRNAs). Remarkably, more than 70% of these changes occurred under the microaerobic conditions typical of symbiotic root nodules. Many SmRNase III-dependent transcripts encode pathways supporting microaerobic metabolism and nitrogen fixation in endosymbiotic bacteroids. Analysis of sequencing read coverage revealed putative consensus cleavage signatures biased toward mRNA 5' untranslated regions, suggesting preferential processing at these sites. Altered expression of sRNAs and/or their predicted mRNA targets further supports a role for SmRNase III in sRNA-mediated silencing. Consistently, in vivo and in vitro assays demonstrated that SmRNase III is required for the repression of nifK (encoding the {beta}-subunit of the nitrogenase MoFe protein) and dctA (encoding a major dicarboxylate transporter) by the antisense sRNA asNifK1 and the trans-sRNA AbcR1, respectively. Our findings reveal a major impact of SmRNase III on shaping the symbiotic transcriptome of S. meliloti and provide a foundation for deeper investigation into the mechanisms and regulatory roles of RNase III activity in rhizobia.