Abstract
The current working model of the innate immune protein calprotectin (CP) focuses on its metal-sequestering activity, which contributes to host defense against infection. Recently, CP was reported to enhance the survival of Staphylococcus aureus in coculture with Pseudomonas aeruginosa in a metal-independent manner. This prior work indicated that the CP protein scaffold, even in the absence of its metal-binding sites, possesses activities that impact interspecies dynamics between these bacterial pathogens. In this study, we employ ΔΔ, a CP variant lacking both functional metal-binding sites, to assess the responses of each pathogen to the CP protein scaffold in monoculture and coculture. Using dual-species transcriptomics, we report that ΔΔ treatment induced gene expression changes indicative of cell envelope modifications for both P. aeruginosa and S. aureus during coculture. The presence of the CP protein scaffold also attenuated the production of the quorum sensing molecule C(4)-homoserine lactone and the anti-staphylococcal alkylquinolone (AQ) metabolite 2-heptyl-4-hydroxyquinoline N-oxide. Cocultures with S. aureus and P. aeruginosa mutants defective in AQ biosynthesis demonstrated that AQ production was required for ΔΔ to impact expression of membrane remodeling genes in both species during coculture. Furthermore, we showed that in the absence of AQ production, the effect of CP on S. aureus in coculture resembled that of Fe depletion. Collectively, our findings demonstrate that the functional versatility of CP extends beyond multi-metal sequestration and that its intertwined metal-dependent and -independent activities have important consequences for bacterial physiology and polymicrobial interactions.