Determinants of metal-specific transcriptional responses in bacteria

Abstract

Metals are essential cofactors for life. However, their beneficial intrinsic chemical properties can be toxic if their cellular levels are not tightly regulated. In a bacterial cell, this control rests with a set of metal-sensing transcriptional regulators that respond to changes in intracellular metal availability to tune the expression of genes required for metal homeostasis. The correct metal ion, bound to its cognate sensor, acts allosterically to alter DNA-binding affinity leading to changes in promoter occupancy. The determinants for a metal-specific response are well understood. However, the problems posed by the mis-metalation of metallosensors exposed to excess non-cognate metal ions have only recently been considered. This study focuses on the response of a set of metalloregulators (from a single organism) to one metal to understand at a molecular level the risk for mis-metalation and, potentially, incorrect function.

The cellular complement of Salmonella enterica serovar Typhimurium metallosensors (MntR, Fur, RcnR, NikR, CueR, ZntR, and Zur) were purified, and their individual affinities for Ni(II), along with the effect of Ni(II) on DNA-binding, were determined using biophysical methods. The two cognate Ni(II)-sensors (NikR and RcnR) showed the tightest Ni(II) affinities. The remaining five sensors had weaker affinities that clustered within a narrow range below a threshold two orders of magnitude weaker than for RcnR. DNA-binding affinities suggest all the non-cognate sensors were capable of a Ni(II)-induced allosteric response in vitro. Simulations of cellular DNA-binding using Ni(II) and DNA affinities with estimates of protein copy number demonstrated that free Ni(II) must be buffered to low levels to ensure the fidelity of Ni(II)-specific transcriptional responses. Fur was identified as the non-cognate sensor most likely to be mis-metalated and mal-respond to Ni(II). The work suggests a general principle for maintaining metal-specific transcriptional responses by buffering individual metal ions within specific ranges.