Anticytolytic Screen Identifies Inhibitors of Mycobacterial Virulence Protein Secretion

Rybniker, Jan; Chen, Jeffrey M.; Sala, Claudia; Hartkoorn, Ruben C.; Vocat, Anthony; Benjak, Andrej; Boy‐Röttger, Stefanie; Zhang, Ming; Székely, Rita; Greff, Zoltán; Őrfi, László; Szabadkai, István; Pató, János; Kéri, Gÿorgý; Cole, Stewart T.

doi:10.1016/j.chom.2014.09.008

Cited by 86 publications

(81 citation statements)

References 36 publications

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“…Additionally, zinc's regulatory effect on bacterial secretion has been demonstrated. Specifically, zinc stress activates the secretion of the EsxA toxin, a substrate of the type VII ESX-1 secretion system in Mycobacterium tuberculosis which arrests phagosome maturation, lyses cell membranes, and induces cell death (73,74). Conversely, zinc negatively regulates the type III secretion system in enteropathogenic E. coli, the ZirTS antivirulence secretion pathway in Salmonella, as well as the type VI secretion system in Burkholderia mallei and Burk-…”

Section: Discussionmentioning

confidence: 99%

The Human Antimicrobial Protein Calgranulin C Participates in Control of Helicobacter pylori Growth and Regulation of Virulence

et al. 2015

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f During infectious processes, antimicrobial proteins are produced by both epithelial cells and innate immune cells. Some of these antimicrobial molecules function by targeting transition metals and sequestering these metals in a process referred to as "nutritional immunity." This chelation strategy ultimately starves invading pathogens, limiting their growth within the vertebrate host. Recent evidence suggests that these metal-binding antimicrobial molecules have the capacity to affect bacterial virulence, including toxin secretion systems. Our previous work showed that the S100A8/S100A9 heterodimer (calprotectin, or calgranulin A/B) binds zinc and represses the elaboration of the H. pylori cag type IV secretion system (T4SS). However, there are several other S100 proteins that are produced in response to infection. We hypothesized that the zinc-binding protein S100A12 (calgranulin C) is induced in response to H. pylori infection and also plays a role in controlling H. pylori growth and virulence. To test this, we analyzed gastric biopsy specimens from H. pylori-positive and -negative patients for S100A12 expression. These assays showed that S100A12 is induced in response to H. pylori infection and inhibits bacterial growth and viability in vitro by binding nutrient zinc. Furthermore, the data establish that the zinc-binding activity of the S100A12 protein represses the activity of the cag T4SS, as evidenced by the gastric cell "hummingbird" phenotype, interleukin 8 (IL-8) secretion, and CagA translocation assays. In addition, high-resolution field emission gun scanning electron microscopy (FEG-SEM) was used to demonstrate that S100A12 represses biogenesis of the cag T4SS. Together with our previous work, these data reveal that multiple S100 proteins can repress the elaboration of an oncogenic bacterial surface organelle. Helicobacter pylori is a Gram-negative member of the Epsilonproteobacteria commonly found in the human stomach (1). More than 50% of the world's population is chronically infected with H. pylori, despite a robust immune response to this bacterium (2, 3). The vast majority of infected persons exhibit no symptoms of disease; however, the presence of this pathogen can increase the risk of negative outcomes, including duodenal ulcer, dysplasia, mucosa-associated lymphoid tissue (MALT) lymphoma, and invasive gastric cancer (4). Negative outcomes of H. pylori infection are associated with multiple factors, including host genetics, environmental contributions such as diet and smoking, and bacterial virulence properties.One major virulence factor that contributes to H. pylori-associated disease outcomes is the cag pathogenicity island-encoded type IV secretion system (cag T4SS) (5). The cag T4SS is a macromolecular nanomachine responsible for translocating substrates, including peptidoglycan and the oncogenic effector molecule CagA, into host epithelial cells (6). The translocation activity of the cag T4SS leads to multiple consequences in host cell biology, including cytoskeletal rearrangements, indu...

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Section: Discussionmentioning

confidence: 99%

The Human Antimicrobial Protein Calgranulin C Participates in Control of Helicobacter pylori Growth and Regulation of Virulence

et al. 2015

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“…To our knowledge, there are no known chemical inhibitors of the PhoPR regulon. Recently, Rybniker and colleagues identified an antivirulence inhibitor of Esx-1 secretion using an anticytolytic screen (31), although the mechanism of this compound appears to be PhoPR independent. ETZ does not alter M. tuberculosis cytoplasmic pH homeostasis (Fig.…”

Section: Discussionmentioning

confidence: 99%

The Carbonic Anhydrase Inhibitor Ethoxzolamide Inhibits the Mycobacterium tuberculosis PhoPR Regulon and Esx-1 Secretion and Attenuates Virulence

Johnson

Colvin

Needle

et al. 2015

Antimicrob Agents Chemother

108

130

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bMycobacterium tuberculosis must sense and adapt to host environmental cues to establish and maintain an infection. The twocomponent regulatory system PhoPR plays a central role in sensing and responding to acidic pH within the macrophage and is required for M. tuberculosis intracellular replication and growth in vivo. Therefore, the isolation of compounds that inhibit PhoPR-dependent adaptation may identify new antivirulence therapies to treat tuberculosis. Here, we report that the carbonic anhydrase inhibitor ethoxzolamide inhibits the PhoPR regulon and reduces pathogen virulence. We show that treatment of M. tuberculosis with ethoxzolamide recapitulates phoPR mutant phenotypes, including downregulation of the core PhoPR regulon, altered accumulation of virulence-associated lipids, and inhibition of Esx-1 protein secretion. Quantitative single-cell imaging of a PhoPR-dependent fluorescent reporter strain demonstrates that ethoxzolamide inhibits PhoPR-regulated genes in infected macrophages and mouse lungs. Moreover, ethoxzolamide reduces M. tuberculosis growth in both macrophages and infected mice. Ethoxzolamide inhibits M. tuberculosis carbonic anhydrase activity, supporting a previously unrecognized link between carbonic anhydrase activity and PhoPR signaling. We propose that ethoxzolamide may be pursued as a new class of antivirulence therapy that functions by modulating expression of the PhoPR regulon and Esx-1-dependent virulence. Mycobacterium tuberculosis is a successful pathogen because it overcomes a variety of obstacles raised by the host immune response. The ability of M. tuberculosis to sense host immune pressures and orchestrate adaptive responses to these cues is essential for pathogen virulence. It follows that the identification of chemical compounds that disrupt the ability of M. tuberculosis to sense and respond to host cues may function to attenuate pathogen virulence.M. tuberculosis uses environmental pH as a cue to modulate its physiology. These pH-dependent adaptations play a central role in M. tuberculosis pathogenesis (1). Transcriptional profiling of M. tuberculosis-infected macrophages identified an M. tuberculosis regulon induced at acidic pH (2) that significantly overlaps the PhoPR two-component regulatory system regulon (3-5), suggesting a role for PhoPR in pH-driven adaptation. phoPR-inactivated M. tuberculosis mutant strains are highly attenuated during mouse and guinea pig infections, supporting that this pathway is a suitable target for new drug development (6, 7).Conventional antimicrobial discovery campaigns seeking to identify bactericidal or bacteriostatic compounds are often performed in vitro. However, many targets required for M. tuberculosis pathogenesis, including environmental sensing pathways, are essential only in vivo and would be missed using standard approaches. Compounds that target in vivo essential bacterial virulence factors are known as antivirulence therapies (8). Antivirulence therapies are advantageous over traditional antibiotics because they preserve e...

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“…46 Intriguingly, another screen for compounds that inhibit the ability of Mtb to kill cultured cells found a small molecule, a benzyloxybenzylidene-hydrazine compound called BBH7, which strongly induces the RicR regulon. 47 BBH7 was selected for analysis because it inhibits the secretion of a major virulence protein called EsxA. 47 Based on these studies, it is tempting to speculate that there is a link between Cu homeostasis and virulence protein secretion in Mtb .…”

Section: Conclusion and Prospectsmentioning

confidence: 99%

Copper homeostasis in Mycobacterium tuberculosis

Shi

Darwin

2015

Metallomics

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Copper (Cu) is a trace element essential for the growth and development of almost all organisms, including bacteria. However, Cu overload in most systems is toxic. Studies show Cu accumulates in macrophage phagosomes infected with bacteria, suggesting Cu provides an innate immune mechanism to combat invading pathogens. To counteract the host-supplied Cu, increasing evidence suggests that bacteria have evolved Cu resistance mechanisms to facilitate their pathogenesis. In particular, Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis, has evolved multiple pathways to respond to Cu. Here, we summarize what is currently known about Cu homeostasis in Mtb and discuss potential sources of Cu encountered by this and other pathogens in a mammalian host.

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Anticytolytic Screen Identifies Inhibitors of Mycobacterial Virulence Protein Secretion

Cited by 86 publications

References 36 publications

The Human Antimicrobial Protein Calgranulin C Participates in Control of Helicobacter pylori Growth and Regulation of Virulence

The Human Antimicrobial Protein Calgranulin C Participates in Control of Helicobacter pylori Growth and Regulation of Virulence

The Carbonic Anhydrase Inhibitor Ethoxzolamide Inhibits the Mycobacterium tuberculosis PhoPR Regulon and Esx-1 Secretion and Attenuates Virulence

Copper homeostasis in Mycobacterium tuberculosis

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