A Redox-Responsive Transcription Factor Is Critical for Pathogenesis and Aerobic Growth of Listeria monocytogenes

Abstract

Bacterial pathogens have evolved sophisticated mechanisms to sense and adapt to redox stress in nature and within the host. However, deciphering the redox environment encountered by intracellular pathogens in the mammalian cytosol is challenging, and that environment remains poorly understood. In this study, we assessed the contributions of the two redox-responsive, Spx-family transcriptional regulators to the virulence of Listeria monocytogenes, a Gram-positive facultative intracellular pathogen. Spx-family proteins are highly conserved in Firmicutes, and the L. monocytogenes genome contains two paralogues, spxA1 and spxA2. Here, we demonstrate that spxA1, but not spxA2, is required for the oxidative stress response and pathogenesis. SpxA1 function appeared to be conserved with the Bacillus subtilis homologue, and resistance to oxidative stress required the canonical CXXC redox-sensing motif. Remarkably, spxA1 was essential for aerobic growth, demonstrating that L. monocytogenes SpxA1 likely regulates a distinct set of genes. Although the ΔspxA1 mutant did not grow in the presence of oxygen in the laboratory, it was able to replicate in macrophages and colonize the spleens, but not the livers, of infected mice. These data suggest that the redox state of bacteria during infection differs significantly from that of bacteria growing in vitro. Further, the host cell cytosol may resemble an anaerobic environment, with tissue-specific variations in redox stress and oxygen concentration.

Inactivation of cysL Inhibits Biofilm Formation by Activating the Disulfide Stress Regulator Spx in Bacillus subtilis

“…Bacterial pathogens have evolved to adapt to their host environment and robustly respond to reactive oxygen species (ROS) generated by host immune cells to kill invading pathogens (38). In many pathogens of Firmicutes, Spx has also evolved to play a critical role in virulence and survival within the host (39)(40)(41)(42)(43)(44). However, my findings indicate that the negative regulation of biofilm formation by Spx is not specific to those pathogens but widespread among many Firmicutes species.…”

YjbH Requires Its Thioredoxin Active Motif for the Nitrosative Stress Response, Cell-to-Cell Spread, and Protein-Protein Interactions in Listeria monocytogenes

“…The only documented role for YjbH in Firmicutes is to modulate levels of Spx such that a strain lacking yjbH has increased Spx abundance (14,16,27) The roles of yjbH Lm and spxA1 were first examined in an SNP MIC assay. An spxA1 knockdown strain (P-spxA1::Tn) that expresses 10-fold less spxA1 transcript was used for these experiments, as a strain deleted for spxA1 does not grow in the presence of oxygen (32). The spxA1 knockdown strain grows aerobically and is more sensitive to hydrogen peroxide and diamide (11), but has not yet been tested in the presence of SNP.…”

“…Instead, YjbH Bs acts as an adaptor by lowering the conformational entropy of Spx Bs , thereby increasing the on September 30, 2020 by guest http://jb.asm.org/ Downloaded from efficiency of Spx Bs binding to ClpX(24). We predict this mechanism is conserved in L. monocytogenes, as SpxA1 shares 83% amino acid identity with Spx Bs and expressing spx Bs functionally complements the ∆spxA1 mutant(32). The second piece of evidence that YjbH Lm is a protease adaptor for SpxA1 comes from whole-cell proteomics, which revealed increased SpxA1 protein in the ∆yjbH Lm strain.…”

Statistical modelling of bacterial promoter sequences for regulatory motif discovery with the help of transcriptome data: application to Listeria monocytogenes

“…In brief, 89 promoter regions are predicted to contain both the -10 and -35 boxes, 88 out of them were previously reported as probable members of the SigB regulon in either [34] or [35]. Spx was identified based on (i) its position with respect to the TSS just upstream of the SigA -35 box, (ii) sequence properties reported for Spx in B. subtilis described as an AGCA element at position -44 [36], and (iii) literature data on the Spx-regulon of L. monocytogenes [37]. PWMs found for PrfA and VirR, two key transcription regulators involved in L. monocytogenes virulence, were in line with previously described sequence properties [38,39].…”