Manganese management

Mol. Cell 57, 1110–1123 (2015)

Riboswitches are RNA motifs that adaptively bind metabolites and couple ligand recognition to regulation of gene expression. Though the three-dimensional structures and the gene-regulatory mechanisms of numerous riboswitch families have been characterized, the physiological ligands of certain riboswitches remain unknown. For instance, the identity of the ligand for the abundant yybP-ykoY riboswitch, which was first described in Bacillus subtilis over a decade ago, has not been established. Now Price et al. provide evidence that the yybP-ykoY riboswitch is a Mn²⁺-responsive transcriptional ON switch. Based on recent data showing that yybP-ykoY is found upstream of genes linked to Mn²⁺ trafficking, the authors hypothesized that the riboswitch may be a Mn²⁺ sensor. In vitro transcription assays with yybP-ykoY from Lactococcus lactis revealed that Mn²⁺, but not other transition-metal ions, relieved basal transcriptional termination, an effect that was confirmed in cells using a reporter gene construct. X-ray crystallographic and mutational analysis alongside chemical probing data of the L. lactis riboswitch revealed a riboswitch architecture based on two coaxially stacked helices organized by a four-way junction that, in the presence of Mn²⁺, creates two phosphate-rich metal-binding sites; one site could be assigned as a Mg²⁺ binding pocket, while the other serves as the Mn²⁺-sensing site and features an octahedral coordination geometry at the metal ion that includes a 'soft' N7 purine ligand donated by A41. Finally, exploring the link between the riboswitch and the physiological effects of elevated Mn²⁺ concentrations, the authors showed that expression of yybP-ykoY–linked YoaB from L. lactis, a putative P-type ATPase, was able to rescue sensitivity to elevated Mn²⁺ concentrations in B. subtilis, providing support for the function of YoaB as a Mn²⁺ efflux pump. Taken together, the combined biochemical and structural data establish Mn²⁺ as the physiological ligand of the yybP-ykoY riboswitch.