Staphylococcus aureus HrtA-heme-toxicity : ATPase function and heme-stress virulence reprogramming in S. aureus Original paper

What was studied?

This study investigates Staphylococcus aureus HrtA-heme-toxicity, defining the biochemical function of HrtA, the ATPase subunit of the heme-regulated transporter HrtAB, and linking its catalytic activity to survival under heme stress and to genome-wide transcriptional remodeling. The authors characterize HrtA ATPase kinetics in vitro, identify essential catalytic residues by site-directed mutagenesis, test in vivo complementation during growth with toxic heme, and profile the heme-stress transcriptome of an hrtA mutant to explain previously observed virulence shifts. The work positions HrtAB at the interface of heme acquisition and heme detoxification in Staphylococcus aureus pathogenesis, a setting relevant to iron competition in host niches and to immunomodulatory toxin expression that shapes host–microbe interactions.

Who was studied?

Experiments used S. aureus strain Newman and an isogenic hrtA deletion mutant for growth and transcriptional analyses, with Escherichia coli BL21(DE3) expressing recombinant HrtA and engineered HrtA mutants for biochemical assays. Heme exposure for microarrays was 1 μM to permit growth of the mutant, while adaptation and dominance tests used up to 10 μM heme; biochemical assays assessed pH, temperature, ATP concentration, and divalent cation requirements. These designs isolated HrtA’s role in heme tolerance and linked catalytic competence to cellular phenotypes and global gene expression under heme stress.

Most important findings

Purified HrtA hydrolyzed ATP, establishing it as the ATPase of the HrtAB transporter. Activity saturated near 0.25 mM ATP, was optimal at pH 7 to 8, and required Mn²⁺ or Mg²⁺, with an unusual in vitro temperature optimum of 10 to 20°C as depicted in Figure 2 on page 4. Conserved motifs were functionally essential: K45 in Walker A, G145 in the ABC signature, and E167 in Walker B were each required for ATP hydrolysis; R76 outside these motifs reduced but did not abolish activity. Only catalytically competent HrtA complemented the heme-sensitive growth defect of the hrtA mutant, whereas catalytically inactive variants acted dominant-negatively in wild-type cells under heme challenge, consistent with multimeric ABC transporter architecture.

Genome-wide expression profiling revealed that more than 500 transcripts changed at least twofold in the hrtA mutant during heme exposure, with the pattern most similar to the stringent response, including approximately 37.4 percent overlap and strong up-regulation of RelA. Virulence determinants were markedly reprogrammed: transcripts for staphylococcal superantigen-like exotoxins were induced, while pore-forming toxins such as gamma hemolysin subunits and a leukotoxin were repressed, indicating a shift from cell-lytic to immunomodulatory toxin profiles. Table 3 on page 7 details these changes, including increases in fibrinogen-binding proteins and immunodominant antigens. Regulatory systems, including HssRS and multiple two-component regulators and metal homeostasis repressors such as Fur and CzrA, were differentially expressed, offering mechanisms for the breadth of the response.

Key implications

For microbiome-oriented pathogenesis frameworks, these data support heme as a contextual host signal that directs S. aureus virulence programming. HrtAB function safeguards against heme’s membrane-damaging toxicity and prevents a costly, immunomodulatory stress response; loss of HrtA precipitates stringent-like reprogramming and favors secreted factors that can reshape immune recruitment in host tissues. In microbiome signatures, S. aureus emerges as a pathobiont whose toxin repertoire and immune interference vary with iron-heme availability, suggesting niche-specific expression states during dysbiosis. Therapeutically, the results raise the prospect of targeting HrtAB or its regulator HssRS to potentiate heme stress and blunt virulence without directly inhibiting growth, a strategy that could complement nutritional immunity and reduce selective pressure for resistance.

Citation

Stauff DL, Bagaley D, Torres VJ, et al. Staphylococcus aureus HrtA Is an ATPase Required for Protection against Heme Toxicity and Prevention of a Transcriptional Heme Stress Response. Journal of Bacteriology. 2008;190(10):3588-3596. doi:10.1128/JB.01921-07