Abstract
ABSTRACTBacteria that colonize animals must overcome, or coexist, with the reactive oxygen species products of inflammation, a front-line defense of innate immunity. Among these is the neutrophilic oxidant bleach, hypochlorous acid (HOCl), a potent antimicrobial that plays a primary role in killing bacteria through nonspecific oxidation of proteins, lipids, and DNA. Here, we report that in response to increasing HOCl levels, Escherichia coli regulates biofilm production via activation of the diguanylate cyclase DgcZ. We identify the mechanism of DgcZ sensing of HOCl to be direct oxidation of its regulatory chemoreceptor zinc-binding (CZB) domain. Dissection of CZB signal transduction reveals that oxidation of the conserved zinc-binding cysteine controls CZB Zn2+ occupancy, which in turn regulates the catalysis of c-di-GMP by the associated GGDEF domain. We find DgcZ-dependent biofilm formation and HOCl sensing to be regulated in vivo by the conserved zinc-coordinating cysteine. Additionally, point mutants that mimic oxidized CZB states increase total biofilm. A survey of bacterial genomes reveals that many pathogenic bacteria that manipulate host inflammation as part of their colonization strategy possess CZB-regulated diguanylate cyclases and chemoreceptors. Our findings suggest that CZB domains are zinc-sensitive regulators that allow host-associated bacteria to perceive host inflammation through reactivity with HOCl.
Highlights
Host-associated bacteria use a repertoire of sensory proteins to eavesdrop on host chemical cues and adapt their lifestyles
No difference in biofilm was observed after 24 h for controls containing untreated cells, for water-treated strains, or for strains treated with phosphate-buffered saline (PBS) buffer at pH 7 (Fig. 1A)
Chemoreceptor zinc-binding (CZB) domains are a class of sensors that have remained enigmatic despite their prevalence among diguanylate cyclases and chemoreceptors in diverse bacterial phyla (Fig. 6) [33]
Summary
Host-associated bacteria use a repertoire of sensory proteins to eavesdrop on host chemical cues and adapt their lifestyles . One of the most consequential host processes for bacteria that colonize animals is inflammation, in which neutrophils infiltrate tissue and generate reactive oxygen and nitrogen species to control and eliminate invading microbes [5, 6] Of these cytotoxic species, hypochlorous acid (HOCl) has been shown to play a dominant role in killing bacteria [7]. Individuals with chronic gastrointestinal inflammation, such as Crohn’s disease and ulcerative colitis, are well known to exhibit disrupted microbiome communities thought to perpetuate a cycle of dysbiosis recalcitrant to correction with therapeutics [31, 32] These examples suggest that inflamed host environments that contain HOCl can dramatically shift the colonization, biogeography, and behavior of host-associated bacteria. It was unclear to what extent our findings on the chemoreceptor TlpD could be extrapolated to the diverse array of CZBcontaining proteins that exist in nature, such as CZB-regulated diguanylate cyclases
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