Bacterial Responses to Iron Withholding by Calprotectin

TL;DR

This perspective reviews recent evidence that human calprotectin (CP) withholds Fe(II) via its His6 site and induces Fe starvation responses in bacterial pathogens including *Pseudomonas aeruginosa*, *Acinetobacter baumannii*, and *Staphylococcus aureus*. It highlights bacterial adaptations such as heme utilization and flavin biosynthesis, and calls for in vivo and environmentally relevant studies of Fe withholding by CP.

Key findings

• CP is described as the first Fe(II)-withholding protein; the Ca(II)-bound heterotetramer binds Fe(II) at its His6 site with subpicomolar affinity, and at near-neutral pH the His6 site relative affinities are Kd,Mn > Kd,Fe > Kd,Zn > Kd,Ni. CP reduces cell-associated Fe in *P. aeruginosa*, *A. baumannii*, *S. aureus*, *E. coli* K-12 and UTI89, *S.* Typhimurium, and *K. pneumoniae*; in *P. aeruginosa*, this reduction requires the His6 site.
• CP induces Fe starvation responses: *P. aeruginosa* increases pyoverdine and pyoverdine uptake proteins, and CP inhibits phenazine production in an Fe-dependent manner; *A. baumannii* upregulates acinetobactin/fimsbactin biosynthesis and transport, *feoAB*, and *fhuE*, while downregulating bacterioferritins; *S. aureus* upregulates *sirA*, *isdC*, *sbnC*, *sirA/B*, *isdABCDEFG*, and *fhuB*, and downregulates a ferritin family protein and L-serine dehydratase Fe-S cluster subunits.
• Bacterial adaptations to CP include heme utilization: heme prevents CP-induced reduction of cell-associated Fe and mitigates Fe starvation responses in *P. aeruginosa* and *S. aureus*. *A. baumannii* upregulates de novo flavin biosynthesis (including RibBX and NusB) under Fe and Zn starvation by CP, and *S. aureus* riboflavin biosynthesis genes *ribH*, *ribBA*, *ribE*, and *ribD* increase ~1.5-fold with CP treatment.
• Effects are organism- and medium-dependent: earlier studies using a 5:1 Mn:Fe medium or TSB-based medium did not observe Fe reduction, while CP reduces Fe in *S. aureus* in LB but not TSB-based medium; CP does not effectively withhold Mn in mildly acidic environments, and all microbiology studies discussed were performed aerobically at near-neutral pH.

Methods

• Design: a perspective/narrative review; no new primary experiments are reported.
• Data or sample: synthesis of cited studies on bacterial pathogens (*P. aeruginosa*, *A. baumannii*, *S. aureus*, *E. coli*, *S.* Typhimurium, *K. pneumoniae*) and murine infection models, including transcriptional, proteomic, metabolite, and cellular metal-level analyses from the referenced literature.
• Main analysis: qualitative comparison across organisms, growth media, CP metal-binding sites, and host models; no primary statistical analysis is reported.

Limitations

• The paper states that most murine model studies focused on Mn and Zn withholding by CP rather than Fe, that Fe withholding by CP has not yet been evaluated in the host context, that murine CP requires further characterization, and that effects under oxygen-limited and acidic conditions remain unknown; the origins of medium dependence are also unclear.
• Evident from the text: as a perspective, it presents no new primary data, and its conclusions rely on cited studies performed under varying media and conditions, with no in vivo Fe-specific experiments described.