Bacterial iron-sulfur cluster sensors in mammalian pathogens

Abstract

Iron-sulfur clusters act as important cofactors for a number of transcriptional regulators in bacteria, including many mammalian pathogens. The sensitivity of iron-sulfur clusters to iron availability, oxygen tension, and reactive oxygen and nitrogen species enables bacteria to use such regulators to adapt their gene expression profiles rapidly in response to changing environmental conditions. In this review, we discuss how the [4Fe-4S] or [2Fe-2S] cluster-containing regulators FNR, Wbl, aconitase, IscR, NsrR, SoxR, and AirSR contribute to bacterial pathogenesis through control of both metabolism and classical virulence factors. In addition, we briefly review mammalian iron homeostasis as well as oxidative/nitrosative stress to provide context for understanding the function of bacterial iron-sulfur cluster sensors in different niches within the host.

Figure 1. The mechanism of oxygen sensing by FNR during Shigella flexneri colonic infection

A model of Mxi-Spa T3SS regulation by FNR in response to the oxygen concentration gradient in the human colon. It has been demonstrated in E. coli that transcription of fnr is constitutive both in the presence and absence of oxygen and the resulting FNR protein is loaded with a [4Fe-4S] cluster via the Isc Fe-S cluster biosynthesis pathway. In S. flexneri, under anaerobic conditions such as the lumen of the colon, the [4Fe-4S] cluster is stable and [4Fe-4S]-FNR represses transcription of spa32 and spa33. Spa32 is essential for proper T3SS function as it mediates the switch between secretion of needle components and effector proteins^-, while Spa33 is an essential component of the T3SS C-ring where it plays a role in recruiting and exporting T3SS-associated proteins. Due to these repressive effects of FNR, the Mxi-Spa T3SS needles are elongated and primed, but S. flexneri is unable to secrete T3SS effector proteins and invade host cells. However, in areas surrounding host colonic epithelial cells, oxygen levels are increased due to diffusion from the capillary network of cell villi. Under these conditions, the [4Fe-4S]-FNR cluster is oxidized to [2Fe-2S] and eventually lost. This leads to accumulation of the inactive form, apo-FNR, which in E. coli is degraded by ClpXP, allowing for derepression of spa32 and spa33^, . Induction of Spa32 and Spa33 leads to a switch from secretion of needle components to effector proteins allowing S. flexneri to invade host colonic epithelial cells.

Figure 2. The mechanism of IscR-dependent Ysc T3SS regulation during enteropathogenic Yersinia infection

A model of IscR control of the Yersinia Ysc T3SS under differing iron availability, oxygen tension, and ROS concentration. Under anaerobic, low ROS conditions where Yersinia is able to obtain iron, such as the gut lumen, transcription of the isc operon should be limited due to sufficient [2Fe-2S] cluster loading onto IscR (holo-IscR), which recognizes a type 1 DNA-binding motif in the isc promoter to repress transcription in a negative feedback loop. Ysc T3SS expression is predicted to be low under such conditions. However, in tissues that are iron-poor (such as the blood), rich in ROS (such as inflamed tissue), or high in oxygen tension, apo-IscR is predicted to accumulate, leading to stimulation of type II motif-containing promoters including the promoter upstream of the gene encoding LcrF, the Ysc T3SS master regulator. This upregulation may allow increased T3SS expression in niches where Y. pseudotuberculosis requires its T3SS to inhibit uptake and killing by phagocytic cells.

Figure 3. The mechanism of S. Typhimurium NsrR-dependent hmp regulation

A model of NsrR control of hmp encoding a NO detoxifying flavohaemoglobin under increasing nitric oxide stress. During growth of S. Typhimurium when concentrations of nitric oxide are minimal, such as in the environment outside of the mammalian host, the NsrR [2Fe-2S] cluster is stable. Under these conditions, [2Fe-2S]-NsrR is a functional DNA-binding protein and represses transcription of hmp encoding a nitric oxide detoxifying flavohaemoglobin. However, when S. Typhimurium is exposed to NO stress, such as that generated by host macrophages where S. Typhimurium survives and proliferates during mammalian infection, the NsrR [2Fe-2S] cluster is nitrosylated. These conditions lead to cluster destabilization and abolish NsrR DNA-binding activity. As such, hmp expression is no longer repressed and the resulting NO detoxification by flavohaemoglobin allows for S. Typhimurium persistence within host macrophages.

Figure 4. The mechanism of SoxR regulation in the family Enterobacteriaceae and P aeruginosa

A model of SoxR regulatory activity in members of the Enterobacteriaceae and P. aeruginosa. (A) SoxR of Enterobacteria coordinates a NO- and superoxide anion-sensitive [2Fe-2S] cluster. During growth under minimal NO and oxidative stress conditions, the SoxR-[2Fe-2S] cluster is in the reduced form (1+) resulting in abolished DNA-binding activity. However, as the concentration of NO and/or superoxide anion increases, such as within the mammalian host, the [2Fe-2S] cluster is oxidized (2+). Under these conditions, SoxR is an active transcription factor and functions solely to upregulate soxS encoding an AraC-type regulator, which then functions to coordinate gene expression to maintain redox homeostasis. (B) The activity of SoxR of P. aeruginosa is slightly different from that described for Enterobacteria. Specifically, the [2Fe-2S] cluster is oxidized is the presence of pyocyanin, which accumulates during stationary phase, and high oxygen concentrations. Additionally, SoxS is absent from P. aeruginosa and other non-Enterobacteria. As such, the oxidized form, SoxR-[2Fe-2S]²⁺ directly regulates a small subset of genes, mexGHI-ompD. Among other proteins, this operon encodes a multidrug efflux pump required for secretion of the quorum sensing quoromones, N-acylhomoserine lactone (AHL) and 2-heptyl-3-hydroxy-4(1H)-quinolone (PQS). As quorum sensing is required for both acute and chronic infections, SoxR plays an important role in P. aeruginosa pathogenesis.