Cathelicidin Peptides Restrict Bacterial Growth via Membrane Perturbation and Induction of Reactive Oxygen Species

ABSTRACT All metazoans produce antimicrobial peptides (AMPs) that have both broad antimicrobial and immunomodulatory activity. Cathelicidins are AMPs that preferentially kill Gram-negative bacteria in vitro, purportedly by assembling into higher-order structures that perforate the membrane. We utili...

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Autores principales:	Dean A. Rowe-Magnus, Adenine Y. Kao, Antonio Cembellin Prieto, Meng Pu, Cheng Kao
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2019
Materias:	cathelicidins ROS antimicrobial peptides biofilms Gram-negative bacteria single-cell Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/94374ec0b27042d6b462a1355df7c3eb
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spelling	oai:doaj.org-article:94374ec0b27042d6b462a1355df7c3eb2021-11-15T15:59:41ZCathelicidin Peptides Restrict Bacterial Growth via Membrane Perturbation and Induction of Reactive Oxygen Species10.1128/mBio.02021-192150-7511https://doaj.org/article/94374ec0b27042d6b462a1355df7c3eb2019-10-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.02021-19https://doaj.org/toc/2150-7511ABSTRACT All metazoans produce antimicrobial peptides (AMPs) that have both broad antimicrobial and immunomodulatory activity. Cathelicidins are AMPs that preferentially kill Gram-negative bacteria in vitro, purportedly by assembling into higher-order structures that perforate the membrane. We utilized high-resolution, single-cell fluorescence microscopy to examine their mechanism of action in real time. Engineered cathelicidins rapidly bound to Gram-negative and Gram-positive cells and penetrated the cytoplasmic membrane. Rapid failure of the peptidoglycan superstructure in regions of active turnover caused leakage of cytoplasmic contents and the formation of membrane-bound blebs. A mutation anticipated to destabilize interactions between cathelicidin subunits had no effect on bactericidal activity, suggesting that cathelicidins have activities beyond perforating the membrane. Nanomolar concentrations of cathelicidins, although not bactericidal, reduced the growth rate of Gram-negative and Gram-positive bacteria. The cells exhibited expression changes in multiple essential processes, including protein synthesis, peptidoglycan biosynthesis, respiration, and the detoxification of reactive oxygen species (ROS). Time-lapse imaging revealed that ROS accumulation preceded bleb formation, and treatments that reduced cellular ROS levels overcame these bactericidal effects. We propose that that the primary effect of cathelicidins is to induce the production of ROS that damage bacterial molecules, leading to slowed growth or cell death. Given their low circulating levels in vivo, AMPs may serve to slow bacterial population expansion so that cellular immunity systems can respond to and battle the infection. IMPORTANCE Antimicrobial peptides (AMPs) are an important part of the mammalian innate immune system in the battle against microbial infection. How AMPs function to control bacteria is not clear, as nearly all activity studies use nonphysiological levels of AMPs. We monitored peptide action in live bacterial cells over short time frames with single-cell resolution and found that the primary effect of cathelicidin peptides is to increase the production of oxidative molecules that cause cellular damage in Gram-positive and Gram-negative bacteria.Dean A. Rowe-MagnusAdenine Y. KaoAntonio Cembellin PrietoMeng PuCheng KaoAmerican Society for MicrobiologyarticlecathelicidinsROSantimicrobial peptidesbiofilmsGram-negative bacteriasingle-cellMicrobiologyQR1-502ENmBio, Vol 10, Iss 5 (2019)
institution	DOAJ
collection	DOAJ
language	EN
topic	cathelicidins ROS antimicrobial peptides biofilms Gram-negative bacteria single-cell Microbiology QR1-502
spellingShingle	cathelicidins ROS antimicrobial peptides biofilms Gram-negative bacteria single-cell Microbiology QR1-502 Dean A. Rowe-Magnus Adenine Y. Kao Antonio Cembellin Prieto Meng Pu Cheng Kao Cathelicidin Peptides Restrict Bacterial Growth via Membrane Perturbation and Induction of Reactive Oxygen Species
description	ABSTRACT All metazoans produce antimicrobial peptides (AMPs) that have both broad antimicrobial and immunomodulatory activity. Cathelicidins are AMPs that preferentially kill Gram-negative bacteria in vitro, purportedly by assembling into higher-order structures that perforate the membrane. We utilized high-resolution, single-cell fluorescence microscopy to examine their mechanism of action in real time. Engineered cathelicidins rapidly bound to Gram-negative and Gram-positive cells and penetrated the cytoplasmic membrane. Rapid failure of the peptidoglycan superstructure in regions of active turnover caused leakage of cytoplasmic contents and the formation of membrane-bound blebs. A mutation anticipated to destabilize interactions between cathelicidin subunits had no effect on bactericidal activity, suggesting that cathelicidins have activities beyond perforating the membrane. Nanomolar concentrations of cathelicidins, although not bactericidal, reduced the growth rate of Gram-negative and Gram-positive bacteria. The cells exhibited expression changes in multiple essential processes, including protein synthesis, peptidoglycan biosynthesis, respiration, and the detoxification of reactive oxygen species (ROS). Time-lapse imaging revealed that ROS accumulation preceded bleb formation, and treatments that reduced cellular ROS levels overcame these bactericidal effects. We propose that that the primary effect of cathelicidins is to induce the production of ROS that damage bacterial molecules, leading to slowed growth or cell death. Given their low circulating levels in vivo, AMPs may serve to slow bacterial population expansion so that cellular immunity systems can respond to and battle the infection. IMPORTANCE Antimicrobial peptides (AMPs) are an important part of the mammalian innate immune system in the battle against microbial infection. How AMPs function to control bacteria is not clear, as nearly all activity studies use nonphysiological levels of AMPs. We monitored peptide action in live bacterial cells over short time frames with single-cell resolution and found that the primary effect of cathelicidin peptides is to increase the production of oxidative molecules that cause cellular damage in Gram-positive and Gram-negative bacteria.
format	article
author	Dean A. Rowe-Magnus Adenine Y. Kao Antonio Cembellin Prieto Meng Pu Cheng Kao
author_facet	Dean A. Rowe-Magnus Adenine Y. Kao Antonio Cembellin Prieto Meng Pu Cheng Kao
author_sort	Dean A. Rowe-Magnus
title	Cathelicidin Peptides Restrict Bacterial Growth via Membrane Perturbation and Induction of Reactive Oxygen Species
title_short	Cathelicidin Peptides Restrict Bacterial Growth via Membrane Perturbation and Induction of Reactive Oxygen Species
title_full	Cathelicidin Peptides Restrict Bacterial Growth via Membrane Perturbation and Induction of Reactive Oxygen Species
title_fullStr	Cathelicidin Peptides Restrict Bacterial Growth via Membrane Perturbation and Induction of Reactive Oxygen Species
title_full_unstemmed	Cathelicidin Peptides Restrict Bacterial Growth via Membrane Perturbation and Induction of Reactive Oxygen Species
title_sort	cathelicidin peptides restrict bacterial growth via membrane perturbation and induction of reactive oxygen species
publisher	American Society for Microbiology
publishDate	2019
url	https://doaj.org/article/94374ec0b27042d6b462a1355df7c3eb
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_version_	1718426983188135936

Cathelicidin Peptides Restrict Bacterial Growth via Membrane Perturbation and Induction of Reactive Oxygen Species

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