The two-component system ColRS discriminates metal signals to regulate virulence and resistance in Pseudomonas aeruginosa.

Two-component systems (TCSs) in Pseudomonas aeruginosa are essential for sensing and responding to diverse environmental cues, including those associated with ion homeostasis and virulence within the host. However, how specific TCSs discriminate between chemically similar metal signals to fine-tune adaptation remains unclear. Here, we identify the TCS ColRS, conserved across the Pseudomonas genus, as a critical sensor that distinguishes zinc (Zn2+) and copper (Cu2+) to drive divergent survival strategies. First, we establish that the response regulator ColR directly binds the promoters of lipid A modification genes (arnB, eptA), the metal responsive regulator czcR, and the alginate and motility regulator Z (amrZ), genetically linking metal sensing to resistance and virulence circuitry. Phenotypically, ColRS translates these inputs into context-dependent states: enhancing polymyxin tolerance and suppressing motility under copper stress, while sustaining type III secretion system (T3SS) virulence and motility under zinc stress. Mechanistically, this discrimination relies on the sensor kinase ColS, which utilizes a non-canonical E96-H105 interface as a specificity filter to recognize Cu2+ (Kd ≈ 1.11 μM) and distinguish it from Zn2+ (Kd ≈ 19.8 μM). Global proteomics confirms this bifurcation, revealing that Zn2+ triggers focused metabolic tuning, whereas Cu2+ induces broad envelope and motility rewiring. Collectively, our findings reveal ColRS as a pivotal molecular hub that decodes the duality of host metal immunity to optimize pathoadaptation.