Combined proteomic and transcriptomic analysis of the response of Bacillus anthracis to oxidative stress

Abstract

The endospore-forming Gram-positive pathogen Bacillus anthracis is responsible for the usually fatal disease, inhalational anthrax. The success of this pathogen is dependent on its ability to subvert elements of the innate immune system of its animal hosts. B. anthracis spores, which are the main infective agent, are engulfed and germinate in patrolling alveolar macrophages. In order for the infection to progress, the resulting vegetative cells must resist the antimicrobial oxidative burst mounted by the host NADPH oxidase complex. The response of B. anthracis to this and other macrophage-related stresses is therefore of major importance to the success of this pathogen, and consequently we have analysed the superoxide and peroxide stress stimulons of B. anthracis strain UM23C1-2 by means of a combined transcriptomics and proteomics approach. The results show distinct patterns of expression in response to paraquat (endogenous superoxide) and hydrogen peroxide stress. While the main response to paraquat is the induction of iron uptake pathways, the response to peroxide predominantly involves the induction of protection and repair mechanisms. Comparisons between the responses of B. anthracis and related soil bacterium, B. subtilis, reveal differences that are likely to be relevant to their respective habitats.

Cellular Iron Distribution in Bacillus anthracis

“…anthracis was cultured to log phase (OD 540 of ϳ0.3) prior to the addition of 800 M PQ, and time-dependent growth was monitored by measuring changes in optical density relative to that of untreated control cultures. This concentration was selected because our previous data showed that it perturbs but does not prevent growth of B. anthracis and because it was the condition that we previously used for a combined proteomic and transcriptomic study (30). The cultures treated with PQ showed a mild impairment of growth, consistent with previous data (30), and were unable to achieve the same stationary-phase density as that of untreated cultures (Fig.…”

“…B. anthracis is predicted to experience significant oxidative stress within the phagolysosome of the host macrophage. Thus, we have been investigating the responses of B. anthracis to such stresses (30,41). Transcriptomic studies have shown the induction of a number of genes encoding putative iron import and siderophore biosynthetic systems during PQ treatment (28,30), which contrasts with the response of E. coli, which represses iron transport in favor of manganese import (3).…”

“…Thus, we have been investigating the responses of B. anthracis to such stresses (30,41). Transcriptomic studies have shown the induction of a number of genes encoding putative iron import and siderophore biosynthetic systems during PQ treatment (28,30), which contrasts with the response of E. coli, which represses iron transport in favor of manganese import (3). In light of our identification of the major cytosolic iron pools, we investigated whether this induced gene expression gave rise to increased iron import and whether iron occupancy of these known iron pools was altered under superoxide stress.…”

“…Our investigations of the response of B. anthracis to oxidative stress (30,41), which relates to the environment predicted to be experienced within the macrophage phagolysosome, led us to investigate the effects of superoxide stress on iron accumulation and its intracellular distribution. Transcriptomic analyses have previously shown an upregulation of transcripts encoding the siderophore biosynthetic enzymes and putative iron transporters in B. anthracis cells in response to the superoxide-generating reagent paraquat (28,30).…”

Study of exogenous oxidative stress response in Escherichia coli, Pseudomonas spp., Bacillus spp., and Salmonella spp.

“…based on the available information about their survival mechanisms has yet to be undertaken (Christman et al, 1985;Inaoka et al, 1990;Farr and Kogoma, 1991;Givskov et al, 1994;Nitta et al, 2000;Sabra et al, 2002;Kabir et al, 2004;Passalacqua et al, 2006Passalacqua et al, , 2007Den Besten et al, 2009;Noor et al, 2009aNoor et al, , 2009bZuber, 2009;Pohl et al, 2011;Huillet et al, 2012;Murata et al, 2012;Den Besten et al, 2013;Nagamitsu et al, 2013). In relation to current studies on bacterial oxidative stress responses, the present study employed preliminary experiments to resolve the impact of increased concentration of the oxidant.…”