Abstract
Transporters of the cation diffusion facilitator (CDF) family form dimers that export transition metals from the cytosol. The opportunistic pathogen Pseudomonas aeruginosa encodes three homologous CDF genes, czcD (PA0397), aitP (PA1297), and yiiP (PA3963). The three proteins are required for virulence in a plant host model. Disruption of the aitP gene leads to higher Fe2+ and Co2+ sensitivity together with an intracellular accumulation of these ions and to a decreased survival in presence of H2O2. Strains lacking czcD and yiiP showed low Zn2+ sensitivity. However, in iron-rich media and in the presence of Zn2+ these strains secreted higher levels of the iron chelator pyoverdine. Disruption of czcD and yiiP in a non-pyoverdine producer strain and lacking the Zn2+-transporting ATPase, increased the Zn2+ sensitivity and the accumulation of this ion. Most importantly, independent of the pyoverdine production strains lacking CzcD or YiiP, presented lower resistance to imipenem, ciprofloxacin, chloramphenicol, and gentamicin. These observations correlated with a lower survival rate upon EDTA-lysozyme treatment and overexpression of OprN and OprD porins. We hypothesize that while AitP is an Fe2+/Co2+ efflux transporter required for Fe2+ homeostasis, and ultimately redox stress handling, CzcD, and YiiP export Zn2+ to the periplasm for proper Zn2+-dependent signaling regulating outer membrane stability and therefore antibiotic tolerance.
Highlights
Transition metals (TM) participate in vital physiological processes such as respiration, reactive oxygen species handling, and transcriptional regulation (Fraústo Da Silva and Williams, 2001)
P. aeruginosa PAO1 cation diffusion facilitator (CDF) transporter sequences were analyzed by multiple sequence alignment including the archetypical Zn/Fe/Cu CDF transporters, YiiP from E. coli and CzcD from Bacillus subtilis (Grass et al, 2005; Moore et al, 2005)
The presence of a cytosolic poly-histidine stretch between transmembrane segments (TMS) 4 and TMS 5 in PA1297/AitP suggests that this CDF is involved in Co2+ fluxes as similar low complexity histidine-rich sequences are found in Co2+ transporting members (Montanini et al, 2007; Podar et al, 2012)
Summary
Transition metals (TM) participate in vital physiological processes such as respiration, reactive oxygen species handling, and transcriptional regulation (Fraústo Da Silva and Williams, 2001). Bacteria face host-driven changes in TM bioavailability, leading to their scarcity or abundance, both conditions detrimental for the bacteria. An excess of these nutrients leads to cellular toxicity, through mechanisms involving the displacement of cognate. CDF Roles during Pseudomonas Virulence metals from catalytic and structural sites of metalloproteins (Ranquet et al, 2007), or by producing reactive oxygen species through Fenton-like reactions (Macomber and Imlay, 2009; Imlay, 2014). A large number of membrane transporter families participate in metal efflux or uptake in order to maintain each individual TM quota and satisfy the metabolic demand
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