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 ( Sm RNase III) in Sinorhizobium meliloti , the symbiotic partner of alfalfa ( Medicago sativa L.). Loss of Sm RNase 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 nodules. Many Sm RNase III-dependent transcripts encode pathways supporting microaerobic metabolism and nitrogen fixation in endosymbiotic bacteroids. Analysis of sequencing read coverage identified putative consensus cleavage signatures enriched in 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 Sm RNase III in sRNA-mediated silencing. Consistently, in vitro assays showed that base‑pairing between nifK (encoding the β‑subunit of the nitrogenase MoFe protein) and the antisense RNA asNifK promotes Sm RNase III-mediated cleavage. In vivo assays further supported that silencing of nifK and dctA (encoding a major dicarboxylate transporter) requires Sm RNase III, with dctA regulation involving a base‑pairing interaction between the trans ‑sRNA AbcR1 and a predicted Sm RNase III cleavage site within the mRNA. Our findings reveal a major impact of Sm RNase III on shaping the symbiotic transcriptome of S. meliloti and provide a foundation for deeper investigation into RNase III-mediated regulation in rhizobia. • S. meliloti RNase III drives RNA homeostasis under microaerobic conditions. • Sm RNase III influences coding and sRNA networks for key symbiotic pathways. • Sm RNase III shapes sRNA-mediated nitrogenase and metabolic regulation.
Guedes-García et al. (Wed,) studied this question.