Proteomics reveal temperature-coupled cobalamin homeostasis and pathogenicity in Pseudomonas aeruginosa

Abstract

The opportunistic pathogen Pseudomonas aeruginosa is highly adaptable to different environmental conditions due to its versatile sensing and metabolic capabilities. Both external temperature and metal availability have a strong influence on the virulence and pathogenicity of P. aeruginosa, but the coupling between these two factors is not well understood. While iron is recognized as major player in nutritional immunity, the role of cobalt and the cobalt-containing vitamin B12 (cobalamin) during host infection remains unclear. Here, we investigate the environmental isolate P. aeruginosa PA254 using high-resolution global proteomics and cellular cobalamin measurements over a temperature gradient spanning environmental, host-associated, and heat-stress conditions (22-42 C). PA254 occupies a continuum between an ambient-temperature state characterized by secreted virulence factors, exopolysaccharide-rich biofilms, and planktonic swimmers and surface swarmers; and a host-associated state characterized by a distinct set of virulence factors, alginate-dominated biofilms, and a strong proportion of surface twitching motility. Pathway analyses indicate a shift toward carbon sparing, energy conservation, redox control, and metabolic maintenance during a host-adapted lifestyle, along with the increased production of alternative iron acquisition strategies relying on heme and siderophores. Proteins of the cobalamin biosynthetic pathway declined significantly above ambient temperatures, despite constant intracellular B12 concentrations and sustained abundance of B12-dependent enzymes across all conditions. This decoupling of biosynthesis from cellular pools implies temperature-dependent changes in prioritization, biosynthetic efficiency, or recycling within B12-dependent processes, while the lack of B12 production at human body temperatures creates avenues for therapeutics interfering with B12 supply. Altogether, this work highlights a gradual rather than stepwise reprogramming of the P. aeruginosa proteome in response to environmental cues, and highlights proteomics as a comprehensive tool to investigate system level mechanisms of challenging pathogens.