Manganese homeostasis and utilization in pathogenic bacteria

Abstract

Manganese (Mn) is a required cofactor for all forms of life. Given the importance of Mn to bacteria, the host has devised strategies to sequester Mn from invaders. In the macrophage phagosome, NRAMP1 removes Mn and other essential metals to starve intracellular pathogens; in the extracellular space, calprotectin chelates Mn and Zn. Calprotectin-mediated Mn sequestration is a newly appreciated host defense mechanism, and recent findings are highlighted herein. In order to acquire Mn when extracellular concentrations are low, bacteria have evolved efficient Mn acquisition systems that are under elegant transcriptional control. To counteract Mn overload, some bacteria possess Mn-specific export systems that are important in vivo, presumably for control of intracellular Mn levels. Mn transporters, their transcriptional regulators and some Mn-requiring enzymes are necessary for virulence of certain bacterial pathogens, as revealed by animal models of infection. Furthermore, Mn is an important facet of the cellular response to oxidative stress, a host antibacterial strategy. The battle for Mn between host and pathogen is now appreciated to be a major determinant of the outcome of infection. In this MicroReview, the contribution of Mn to the host-pathogen interaction is reviewed, and key questions are proposed for future study.

Figure 1. The battle for Mn at the pathogen-host interface

(A) Intracellular pathogens such as Salmonella enterica sv. Typhimurium that reside within the phagosome of macrophages must survive a Mn deplete environment. Mn and Fe are actively removed from the phagosome by the eukaryotic NRAMP1 protein. Host processes to restrict microbial access to Mn within other cellular compartments, including the cytoplasm, have not yet been identified. To acquire Mn in this restricted environment, Salmonella utilizes MntH and SitABCD, an ABC-family transporter, to acquire Mn and possibly Fe. MntH consists of a single integral membrane protein, whereas ABC-type transporters are made up of a substrate-binding domain, a permease domain, and an ATP-hydrolyzing domain. (B) Extracellular Mn is sequestered by calprotectin, which is released by neutrophils actively or through cell lysis. Other proteins, including lactoferrin and transferrin, have been shown to bind to Mn in plasma or other tissues, but whether Mn-binding by these or as-yet identified extracellular chelators contributes to host nutritional immunity is unknown. To combat this sequestration, extracellular Staphylococcus aureus also utilizes MntH and ABC-type Mn importers. (C) In contrast, Neisseria meningitidis requires Mn export through MntX for virulence, presumably to prevent Mn toxicity during infection. Host mechanisms to intoxicate bacteria with Mn have not yet been identified.

Figure 2. Intracellular utilization of Mnin the oxidative stress response and DNA synthesis

(1) In the presence of hydrogen peroxide, the transcriptional regulator OxyR is oxidized to form an intramolecular disulfide bind that alters the binding affinity of OxyR to the mntH promoter, leading to the activation of mntH transcription. (2) mntH transcription upregulates MntH expression at the cell membrane, increasing Mn import into the cytoplasm where it can be utilized in various processes. (3) Fe-mononuclear enzymes are inactivated by hydrogen peroxide via oxidation of Fe(II) to Fe(III) and dissolution of Fe(III) from the enzyme; Zn can then be incorporated into the enzyme but does not confer enzymatic activity. In the presence of increased Mn either through supplementation or MntH-mediated import, Mn can be incorporated into the mononuclear enzyme and restore enzymatic activity while remaining nonsusceptible to hydrogen peroxide-mediated oxidation. (4) Superoxide anion is dismutased to hydrogen peroxide through the activity of Mn-superoxide dismutase (SOD). (5) Low molecular weight Mn complexes (LMW:Mn), such as Mn-phosphate and Mn-lactate, are capable of detoxifying reactive oxygen species (ROS) in a protein-independent manner. (6) Dps is an Fe-storage protein that protects against hydrogen peroxide through several Mn-requiring activities. First, when Mn is bound to a regulatory site near the N-terminus of the protein, Dps binds to and protects DNA from oxidative damage. Second, Dps ferroxidase activity oxidizes Fe(II) to Fe(III) at the metal-binding site B while reducing hydrogen peroxide, avoiding toxic Fenton chemistry. The ferroxidase activity requires redox cycling of Mn at metal-binding site A. (7) Ribonucleotide reductase NrdEF utilizes a Mn cofactor to reduce ribonucleotides (NTPs) to deoxyribonucleotides (dNTPs). The active cofactor form (Mn(III)-thiyl radical) is derived by the flavodoxin NrdI.

Figure 3. Regulation of Mn import

(A) PerR-mediated derepression of mnt expression is dependent on metal cofactor binding. PerR:Fe is highly susceptible to peroxide-induced oxidation of one of two histidine residues within PerR, which causes PerR to release from the mnt promoter. However, PerR:Mn is not susceptible to oxidation and remains bound. Presumably, when the Mn:Fe ratio is high enough for Mn to populate the PerR regulatory site, Mn levels are also sufficiently high to protect the cell from peroxide stress. (B) MntR and Fur both function as metal-dependent repressors of Mn transport genes in some bacteria. MntR:Mn represses transcription when Mn levels are adequate, but repression is relieved when Mn levels are not sufficiently for MntR to be bound to Mn. Fur can repress Mn transport genes either when bound to Fe or Mn.