A Small-Molecule Modulator of Metal Homeostasis in Gram-Positive Pathogens

ABSTRACT Metals are essential nutrients that all living organisms acquire from their environment. While metals are necessary for life, excess metal uptake can be toxic; therefore, intracellular metal levels are tightly regulated in bacterial cells. Staphylococcus aureus, a Gram-positive bacterium, r...

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Autores principales:	Lillian J. Juttukonda, William N. Beavers, Daisy Unsihuay, Kwangho Kim, Gleb Pishchany, Kyle J. Horning, Andy Weiss, Hassan Al-Tameemi, Jeffrey M. Boyd, Gary A. Sulikowski, Aaron B. Bowman, Eric P. Skaar
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2020
Materias:	MRSA Staphylococcus aureus antibiotics cobalt copper manganese Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/1260e5a12a7c4059be846aa784f59aa4
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spelling	oai:doaj.org-article:1260e5a12a7c4059be846aa784f59aa42021-11-15T16:19:09ZA Small-Molecule Modulator of Metal Homeostasis in Gram-Positive Pathogens10.1128/mBio.02555-202150-7511https://doaj.org/article/1260e5a12a7c4059be846aa784f59aa42020-10-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.02555-20https://doaj.org/toc/2150-7511ABSTRACT Metals are essential nutrients that all living organisms acquire from their environment. While metals are necessary for life, excess metal uptake can be toxic; therefore, intracellular metal levels are tightly regulated in bacterial cells. Staphylococcus aureus, a Gram-positive bacterium, relies on metal uptake and metabolism to colonize vertebrates. Thus, we hypothesized that an expanded understanding of metal homeostasis in S. aureus will lead to the discovery of pathways that can be targeted with future antimicrobials. We sought to identify small molecules that inhibit S. aureus growth in a metal-dependent manner as a strategy to uncover pathways that maintain metal homeostasis. Here, we demonstrate that VU0026921 kills S. aureus through disruption of metal homeostasis. VU0026921 activity was characterized through cell culture assays, transcriptional sequencing, compound structure-activity relationship, reactive oxygen species (ROS) generation assays, metal binding assays, and metal level analyses. VU0026921 disrupts metal homeostasis in S. aureus, increasing intracellular accumulation of metals and leading to toxicity through mismetalation of enzymes, generation of reactive oxygen species, or disruption of other cellular processes. Antioxidants partially protect S. aureus from VU0026921 killing, emphasizing the role of reactive oxygen species in the mechanism of killing, but VU0026921 also kills S. aureus anaerobically, indicating that the observed toxicity is not solely oxygen dependent. VU0026921 disrupts metal homeostasis in multiple Gram-positive bacteria, leading to increased reactive oxygen species and cell death, demonstrating the broad applicability of these findings. Further, this study validates VU0026921 as a probe to further decipher mechanisms required to maintain metal homeostasis in Gram-positive bacteria. IMPORTANCE Staphylococcus aureus is a leading agent of antibiotic-resistant bacterial infections in the world. S. aureus tightly controls metal homeostasis during infection, and disruption of metal uptake systems impairs staphylococcal virulence. We identified small molecules that interfere with metal handling in S. aureus to develop chemical probes to investigate metallobiology in this organism. Compound VU0026921 was identified as a small molecule that kills S. aureus both aerobically and anaerobically. The activity of VU0026921 is modulated by metal supplementation, is enhanced by genetic inactivation of Mn homeostasis genes, and correlates with increased cellular reactive oxygen species. Treatment with VU0026921 causes accumulation of multiple metals within S. aureus cells and concomitant upregulation of genes involved in metal detoxification. This work defines a small-molecule probe for further defining the role of metal toxicity in S. aureus and validates future antibiotic development targeting metal toxicity pathways.Lillian J. JuttukondaWilliam N. BeaversDaisy UnsihuayKwangho KimGleb PishchanyKyle J. HorningAndy WeissHassan Al-TameemiJeffrey M. BoydGary A. SulikowskiAaron B. BowmanEric P. SkaarAmerican Society for MicrobiologyarticleMRSAStaphylococcus aureusantibioticscobaltcoppermanganeseMicrobiologyQR1-502ENmBio, Vol 11, Iss 5 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	MRSA Staphylococcus aureus antibiotics cobalt copper manganese Microbiology QR1-502
spellingShingle	MRSA Staphylococcus aureus antibiotics cobalt copper manganese Microbiology QR1-502 Lillian J. Juttukonda William N. Beavers Daisy Unsihuay Kwangho Kim Gleb Pishchany Kyle J. Horning Andy Weiss Hassan Al-Tameemi Jeffrey M. Boyd Gary A. Sulikowski Aaron B. Bowman Eric P. Skaar A Small-Molecule Modulator of Metal Homeostasis in Gram-Positive Pathogens
description	ABSTRACT Metals are essential nutrients that all living organisms acquire from their environment. While metals are necessary for life, excess metal uptake can be toxic; therefore, intracellular metal levels are tightly regulated in bacterial cells. Staphylococcus aureus, a Gram-positive bacterium, relies on metal uptake and metabolism to colonize vertebrates. Thus, we hypothesized that an expanded understanding of metal homeostasis in S. aureus will lead to the discovery of pathways that can be targeted with future antimicrobials. We sought to identify small molecules that inhibit S. aureus growth in a metal-dependent manner as a strategy to uncover pathways that maintain metal homeostasis. Here, we demonstrate that VU0026921 kills S. aureus through disruption of metal homeostasis. VU0026921 activity was characterized through cell culture assays, transcriptional sequencing, compound structure-activity relationship, reactive oxygen species (ROS) generation assays, metal binding assays, and metal level analyses. VU0026921 disrupts metal homeostasis in S. aureus, increasing intracellular accumulation of metals and leading to toxicity through mismetalation of enzymes, generation of reactive oxygen species, or disruption of other cellular processes. Antioxidants partially protect S. aureus from VU0026921 killing, emphasizing the role of reactive oxygen species in the mechanism of killing, but VU0026921 also kills S. aureus anaerobically, indicating that the observed toxicity is not solely oxygen dependent. VU0026921 disrupts metal homeostasis in multiple Gram-positive bacteria, leading to increased reactive oxygen species and cell death, demonstrating the broad applicability of these findings. Further, this study validates VU0026921 as a probe to further decipher mechanisms required to maintain metal homeostasis in Gram-positive bacteria. IMPORTANCE Staphylococcus aureus is a leading agent of antibiotic-resistant bacterial infections in the world. S. aureus tightly controls metal homeostasis during infection, and disruption of metal uptake systems impairs staphylococcal virulence. We identified small molecules that interfere with metal handling in S. aureus to develop chemical probes to investigate metallobiology in this organism. Compound VU0026921 was identified as a small molecule that kills S. aureus both aerobically and anaerobically. The activity of VU0026921 is modulated by metal supplementation, is enhanced by genetic inactivation of Mn homeostasis genes, and correlates with increased cellular reactive oxygen species. Treatment with VU0026921 causes accumulation of multiple metals within S. aureus cells and concomitant upregulation of genes involved in metal detoxification. This work defines a small-molecule probe for further defining the role of metal toxicity in S. aureus and validates future antibiotic development targeting metal toxicity pathways.
format	article
author	Lillian J. Juttukonda William N. Beavers Daisy Unsihuay Kwangho Kim Gleb Pishchany Kyle J. Horning Andy Weiss Hassan Al-Tameemi Jeffrey M. Boyd Gary A. Sulikowski Aaron B. Bowman Eric P. Skaar
author_facet	Lillian J. Juttukonda William N. Beavers Daisy Unsihuay Kwangho Kim Gleb Pishchany Kyle J. Horning Andy Weiss Hassan Al-Tameemi Jeffrey M. Boyd Gary A. Sulikowski Aaron B. Bowman Eric P. Skaar
author_sort	Lillian J. Juttukonda
title	A Small-Molecule Modulator of Metal Homeostasis in Gram-Positive Pathogens
title_short	A Small-Molecule Modulator of Metal Homeostasis in Gram-Positive Pathogens
title_full	A Small-Molecule Modulator of Metal Homeostasis in Gram-Positive Pathogens
title_fullStr	A Small-Molecule Modulator of Metal Homeostasis in Gram-Positive Pathogens
title_full_unstemmed	A Small-Molecule Modulator of Metal Homeostasis in Gram-Positive Pathogens
title_sort	small-molecule modulator of metal homeostasis in gram-positive pathogens
publisher	American Society for Microbiology
publishDate	2020
url	https://doaj.org/article/1260e5a12a7c4059be846aa784f59aa4
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A Small-Molecule Modulator of Metal Homeostasis in Gram-Positive Pathogens

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