Signaling by the heavy-metal sensor CusS involves rearranged helical interactions in specific transmembrane regions

Abstract

Two-component systems (TCSs) play important roles in the adaptation of bacteria to stress. Despite their increasingly well understood mechanistic features, it remains poorly understood how TCSs transduce signals across membranes. Here, we use the E. coli Cu/Ag-responsive CusSR TCS as a model to investigate the roles of CusS transmembrane (TM) residues. Proline scanning of TM1 domain led to identification of the T17P, F18P, and S21P variants, which display higher kinase activities relative to wild type. A single point mutation, V202G, in the adjacent TM2 domain specifically suppresses the hyperactivities of these mutants. Disulfide crosslinking analysis demonstrated that T17 and V202 are situated in close proximity, and Cys residues substituted at those two positions form exclusive intramolecular crosslinks when CusS is in the signaling-inactive state. In the signaling-active variant of CusS, however, only intermolecular crosslinking between the two Cys residues could be observed, suggesting that destabilization of an intramolecular constraint and a subsequent rearrangement of helical interactions in this TM region is involved in the activation of CusS. An analogous TM helical interface in the P. aeruginosa heavy metal sensor kinase CzcS is also observed. Together, these results suggested a conserved transmembrane signal transduction mechanism in the heavy metal sensing TCSs. Signal transduction of bacterial two-component systems (TCSs) depends on the proper coupling of signal sensing to response output across biological membranes. Here we show that activation of the CusS sensor histidine kinase (HK) of the CusSR TCS is mediated by the dissociation of a transmembrane (TM) intra-molecular constraint containing T17 in TM1 and V202 in TM2. Conservancy of this TM helical interface suggested that it represents a common cross-membrane signal transduction mechanism in the heavy metal sensing TCSs.