Bacillus subtilis TerC Family Proteins Help Prevent Manganese Intoxication

Manganese (Mn) is an essential element and is required for the virulence of many pathogens. In Bacillus subtilis, Mn(II) homeostasis is regulated by MntR, a Mn(II)-responsive, DNA-binding protein. MntR serves as both a repressor of Mn(II) uptake transporters and as a transcriptional activator for expression of two cation diffusion facilitator Mn(II) efflux pumps, MneP and MneS. Mutants lacking either mntR or both mneP and mneS are extremely sensitive to Mn(II) intoxication. Using transposon mutagenesis to select suppressors of Mn(II) sensitivity, we identified YceF, a TerC family membrane protein, as capable of providing Mn(II) resistance. Another TerC paralog, YkoY, is regulated by a Mn(II)-sensing riboswitch and is partially redundant in function with YceF. YkoY is regulated in parallel with an unknown function protein YybP, also controlled by a Mn(II)-sensing riboswitch. Strains lacking between one and five of these known or putative Mn(II) tolerance proteins (MneP, MneS, YceF, YkoY, and YybP) were tested for sensitivity to Mn(II) in growth assays and for accumulation of Mn(II) using inductively coupled plasma mass spectrometry. Loss of YceF and, to a lesser extent, YkoY, sensitizes cells lacking the MneP and MneS efflux transporters to Mn(II) intoxication. This sensitivity correlates with elevated intracellular Mn(II), consistent with the suggestion that TerC proteins function in Mn(II) efflux.IMPORTANCE Manganese homeostasis is primarily regulated at the level of transport. Bacillus subtilis MntR serves as a Mn(II)-activated repressor of importer genes (mntH and mntABC) and an activator of efflux genes (mneP and mneS). Elevated intracellular Mn(II) also binds to Mn-sensing riboswitches to activate transcription of yybP and ykoY, which encodes a TerC family member. Here, we demonstrate that two TerC family proteins, YceF and YkoY, help prevent Mn(II) intoxication. TerC family proteins are widespread in bacteria and may influence host-pathogen interactions, but their effects on Mn(II) homeostasis are unclear. Our results suggest that TerC proteins work by Mn(II) export under Mn(II) overload conditions to help alleviate toxicity.