Genetic and Chemical-Genetic Interactions Map Biogenesis and Permeability Determinants of the Outer Membrane of <named-content content-type="genus-species">Escherichia coli</named-content>

ABSTRACT Gram-negative bacteria are intrinsically resistant to many antibiotics due to their outer membrane barrier. Although the outer membrane has been studied for decades, there is much to uncover about the biology and permeability of this complex structure. Investigating synthetic genetic intera...

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Autores principales:	Kristina Klobucar, Shawn French, Jean-Philippe Côté, James R. Howes, Eric D. Brown
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2020
Materias:	enterobacterial common antigen genetic interaction knockout lipopolysaccharide outer membrane permeability Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/de218b6e7e65427e816e9548a011720d
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spelling	oai:doaj.org-article:de218b6e7e65427e816e9548a011720d2021-11-15T15:57:03ZGenetic and Chemical-Genetic Interactions Map Biogenesis and Permeability Determinants of the Outer Membrane of <named-content content-type="genus-species">Escherichia coli</named-content>10.1128/mBio.00161-202150-7511https://doaj.org/article/de218b6e7e65427e816e9548a011720d2020-04-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00161-20https://doaj.org/toc/2150-7511ABSTRACT Gram-negative bacteria are intrinsically resistant to many antibiotics due to their outer membrane barrier. Although the outer membrane has been studied for decades, there is much to uncover about the biology and permeability of this complex structure. Investigating synthetic genetic interactions can reveal a great deal of information about genetic function and pathway interconnectivity. Here, we performed synthetic genetic arrays (SGAs) in Escherichia coli by crossing a subset of gene deletion strains implicated in outer membrane permeability with nonessential gene and small RNA (sRNA) deletion collections. Some 155,400 double-deletion strains were grown on rich microbiological medium with and without subinhibitory concentrations of two antibiotics excluded by the outer membrane, vancomycin and rifampin, to probe both genetic interactions and permeability. The genetic interactions of interest were synthetic sick or lethal (SSL) gene deletions that were detrimental to the cell in combination but had a negligible impact on viability individually. On average, there were ∼30, ∼36, and ∼40 SSL interactions per gene under no-drug, rifampin, and vancomycin conditions, respectively; however, many of these involved frequent interactors. Our data sets have been compiled into an interactive database called the Outer Membrane Interaction (OMI) Explorer, where genetic interactions can be searched, visualized across the genome, compared between conditions, and enriched for gene ontology (GO) terms. A set of SSL interactions revealed connectivity and permeability links between enterobacterial common antigen (ECA) and lipopolysaccharide (LPS) of the outer membrane. This data set provides a novel platform to generate hypotheses about outer membrane biology and permeability. IMPORTANCE Gram-negative bacteria are a major concern for public health, particularly due to the rise of antibiotic resistance. It is important to understand the biology and permeability of the outer membrane of these bacteria in order to increase the efficacy of antibiotics that have difficulty penetrating this structure. Here, we studied the genetic interactions of a subset of outer membrane-related gene deletions in the model Gram-negative bacterium E. coli. We systematically combined these mutants with 3,985 nonessential gene and small RNA deletion mutations in the genome. We examined the viability of these double-deletion strains and probed their permeability characteristics using two antibiotics that have difficulty crossing the outer membrane barrier. An understanding of the genetic basis for outer membrane integrity can assist in the development of new antibiotics with favorable permeability properties and the discovery of compounds capable of increasing outer membrane permeability to enhance the activity of existing antibiotics.Kristina KlobucarShawn FrenchJean-Philippe CôtéJames R. HowesEric D. BrownAmerican Society for Microbiologyarticleenterobacterial common antigengenetic interactionknockoutlipopolysaccharideouter membranepermeabilityMicrobiologyQR1-502ENmBio, Vol 11, Iss 2 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	enterobacterial common antigen genetic interaction knockout lipopolysaccharide outer membrane permeability Microbiology QR1-502
spellingShingle	enterobacterial common antigen genetic interaction knockout lipopolysaccharide outer membrane permeability Microbiology QR1-502 Kristina Klobucar Shawn French Jean-Philippe Côté James R. Howes Eric D. Brown Genetic and Chemical-Genetic Interactions Map Biogenesis and Permeability Determinants of the Outer Membrane of <named-content content-type="genus-species">Escherichia coli</named-content>
description	ABSTRACT Gram-negative bacteria are intrinsically resistant to many antibiotics due to their outer membrane barrier. Although the outer membrane has been studied for decades, there is much to uncover about the biology and permeability of this complex structure. Investigating synthetic genetic interactions can reveal a great deal of information about genetic function and pathway interconnectivity. Here, we performed synthetic genetic arrays (SGAs) in Escherichia coli by crossing a subset of gene deletion strains implicated in outer membrane permeability with nonessential gene and small RNA (sRNA) deletion collections. Some 155,400 double-deletion strains were grown on rich microbiological medium with and without subinhibitory concentrations of two antibiotics excluded by the outer membrane, vancomycin and rifampin, to probe both genetic interactions and permeability. The genetic interactions of interest were synthetic sick or lethal (SSL) gene deletions that were detrimental to the cell in combination but had a negligible impact on viability individually. On average, there were ∼30, ∼36, and ∼40 SSL interactions per gene under no-drug, rifampin, and vancomycin conditions, respectively; however, many of these involved frequent interactors. Our data sets have been compiled into an interactive database called the Outer Membrane Interaction (OMI) Explorer, where genetic interactions can be searched, visualized across the genome, compared between conditions, and enriched for gene ontology (GO) terms. A set of SSL interactions revealed connectivity and permeability links between enterobacterial common antigen (ECA) and lipopolysaccharide (LPS) of the outer membrane. This data set provides a novel platform to generate hypotheses about outer membrane biology and permeability. IMPORTANCE Gram-negative bacteria are a major concern for public health, particularly due to the rise of antibiotic resistance. It is important to understand the biology and permeability of the outer membrane of these bacteria in order to increase the efficacy of antibiotics that have difficulty penetrating this structure. Here, we studied the genetic interactions of a subset of outer membrane-related gene deletions in the model Gram-negative bacterium E. coli. We systematically combined these mutants with 3,985 nonessential gene and small RNA deletion mutations in the genome. We examined the viability of these double-deletion strains and probed their permeability characteristics using two antibiotics that have difficulty crossing the outer membrane barrier. An understanding of the genetic basis for outer membrane integrity can assist in the development of new antibiotics with favorable permeability properties and the discovery of compounds capable of increasing outer membrane permeability to enhance the activity of existing antibiotics.
format	article
author	Kristina Klobucar Shawn French Jean-Philippe Côté James R. Howes Eric D. Brown
author_facet	Kristina Klobucar Shawn French Jean-Philippe Côté James R. Howes Eric D. Brown
author_sort	Kristina Klobucar
title	Genetic and Chemical-Genetic Interactions Map Biogenesis and Permeability Determinants of the Outer Membrane of <named-content content-type="genus-species">Escherichia coli</named-content>
title_short	Genetic and Chemical-Genetic Interactions Map Biogenesis and Permeability Determinants of the Outer Membrane of <named-content content-type="genus-species">Escherichia coli</named-content>
title_full	Genetic and Chemical-Genetic Interactions Map Biogenesis and Permeability Determinants of the Outer Membrane of <named-content content-type="genus-species">Escherichia coli</named-content>
title_fullStr	Genetic and Chemical-Genetic Interactions Map Biogenesis and Permeability Determinants of the Outer Membrane of <named-content content-type="genus-species">Escherichia coli</named-content>
title_full_unstemmed	Genetic and Chemical-Genetic Interactions Map Biogenesis and Permeability Determinants of the Outer Membrane of <named-content content-type="genus-species">Escherichia coli</named-content>
title_sort	genetic and chemical-genetic interactions map biogenesis and permeability determinants of the outer membrane of <named-content content-type="genus-species">escherichia coli</named-content>
publisher	American Society for Microbiology
publishDate	2020
url	https://doaj.org/article/de218b6e7e65427e816e9548a011720d
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Genetic and Chemical-Genetic Interactions Map Biogenesis and Permeability Determinants of the Outer Membrane of <named-content content-type="genus-species">Escherichia coli</named-content>

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