Antisense transcription of symbiotic genes in Sinorhizobium meliloti

Transcripts that do not encode proteins (ncRNAs) have emerged as a widespread type of regulators in all kingdoms of life. They usually fine-tune gene expression in response to environmental changes by modulating mRNA translation and/or stability through base pairing. Given that bacterial adaptation during the transition from free-living to host-dependent conditions demands a tightly and coordinated regulation of gene expression upon perception of a variety of signals, symbioses, as well as pathogenesis, present a set of characteristics expected to be modulated by ncRNAs. However, little is known about the relevance of ncRNAs in the biology of nitrogen-fixing endosymbiotic bacteria. Here, we mined published RNAseq data from the legume symbiont Sinorhizobium meliloti to search for ncRNAs transcribed opposite to the coding strand (antisense ncRNAs; asRNAs) of genuine symbiotic genes, i.e. required for the synthesis of nodulation signal factors (nod genes) and the nitrogenase components (nif genes). Further characterization of selected nodD2, nifK and nifE asRNAs revealed that some transcripts accumulate differentially in response to different environmental factors, and seem to repress nitrogen fixation genes under conditions in which functional nitrogenase is dispensable, suggesting regulatory functions. Furthermore, Medicago sativa plants inoculated with genetically modified S. meliloti strains overexpressing one of these asRNAs showed slight differences in nitrogen fixation ability with respect to the controls. These experiments provide the first evidences of symbiotically relevant antisense riboregulation in rhizobia.