Loss of <italic toggle="yes">O</italic>-Linked Protein Glycosylation in <named-content content-type="genus-species">Burkholderia cenocepacia</named-content> Impairs Biofilm Formation and Siderophore Activity and Alters Transcriptional Regulators

ABSTRACT O-linked protein glycosylation is a conserved feature of the Burkholderia genus. The addition of the trisaccharide β-Gal-(1,3)-α-GalNAc-(1,3)-β-GalNAc to membrane exported proteins in Burkholderia cenocepacia is required for bacterial fitness and resistance to environmental stress. However,...

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Autores principales:	Cameron C. Oppy, Leila Jebeli, Miku Kuba, Clare V. Oates, Richard Strugnell, Laura E. Edgington-Mitchell, Miguel A. Valvano, Elizabeth L. Hartland, Hayley J. Newton, Nichollas E. Scott
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2019
Materias:	glycosylation pathogenesis Burkholderia cenocepacia posttranslational modifications proteomics DNA binding Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/420dcd1484ec4328b4745ae031fe35ca
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spelling	oai:doaj.org-article:420dcd1484ec4328b4745ae031fe35ca2021-11-15T15:22:24ZLoss of <italic toggle="yes">O</italic>-Linked Protein Glycosylation in <named-content content-type="genus-species">Burkholderia cenocepacia</named-content> Impairs Biofilm Formation and Siderophore Activity and Alters Transcriptional Regulators10.1128/mSphere.00660-192379-5042https://doaj.org/article/420dcd1484ec4328b4745ae031fe35ca2019-12-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mSphere.00660-19https://doaj.org/toc/2379-5042ABSTRACT O-linked protein glycosylation is a conserved feature of the Burkholderia genus. The addition of the trisaccharide β-Gal-(1,3)-α-GalNAc-(1,3)-β-GalNAc to membrane exported proteins in Burkholderia cenocepacia is required for bacterial fitness and resistance to environmental stress. However, the underlying causes of the defects observed in the absence of glycosylation are unclear. Using proteomics, luciferase reporter assays, and DNA cross-linking, we demonstrate the loss of glycosylation leads to changes in transcriptional regulation of multiple proteins, including the repression of the master quorum CepR/I. These proteomic and transcriptional alterations lead to the abolition of biofilm formation and defects in siderophore activity. Surprisingly, the abundance of most of the known glycosylated proteins did not significantly change in the glycosylation-defective mutants, except for BCAL1086 and BCAL2974, which were found in reduced amounts, suggesting they could be degraded. However, the loss of these two proteins was not responsible for driving the proteomic alterations, biofilm formation, or siderophore activity. Together, our results show that loss of glycosylation in B. cenocepacia results in a global cell reprogramming via alteration of the transcriptional regulatory systems, which cannot be explained by the abundance changes in known B. cenocepacia glycoproteins. IMPORTANCE Protein glycosylation is increasingly recognized as a common posttranslational protein modification in bacterial species. Despite this commonality, our understanding of the role of most glycosylation systems in bacterial physiology and pathogenesis is incomplete. In this work, we investigated the effect of the disruption of O-linked glycosylation in the opportunistic pathogen Burkholderia cenocepacia using a combination of proteomic, molecular, and phenotypic assays. We find that in contrast to recent findings on the N-linked glycosylation systems of Campylobacter jejuni, O-linked glycosylation does not appear to play a role in proteome stabilization of most glycoproteins. Our results reveal that loss of glycosylation in B. cenocepacia strains leads to global proteome and transcriptional changes, including the repression of the quorum-sensing regulator cepR (BCAM1868) gene. These alterations lead to dramatic phenotypic changes in glycosylation-null strains, which are paralleled by both global proteomic and transcriptional alterations, which do not appear to directly result from the loss of glycosylation per se. This research unravels the pleiotropic effects of O-linked glycosylation in B. cenocepacia, demonstrating that its loss does not simply affect the stability of the glycoproteome, but also interferes with transcription and the broader proteome.Cameron C. OppyLeila JebeliMiku KubaClare V. OatesRichard StrugnellLaura E. Edgington-MitchellMiguel A. ValvanoElizabeth L. HartlandHayley J. NewtonNichollas E. ScottAmerican Society for MicrobiologyarticleglycosylationpathogenesisBurkholderia cenocepaciaposttranslational modificationsproteomicsDNA bindingMicrobiologyQR1-502ENmSphere, Vol 4, Iss 6 (2019)
institution	DOAJ
collection	DOAJ
language	EN
topic	glycosylation pathogenesis Burkholderia cenocepacia posttranslational modifications proteomics DNA binding Microbiology QR1-502
spellingShingle	glycosylation pathogenesis Burkholderia cenocepacia posttranslational modifications proteomics DNA binding Microbiology QR1-502 Cameron C. Oppy Leila Jebeli Miku Kuba Clare V. Oates Richard Strugnell Laura E. Edgington-Mitchell Miguel A. Valvano Elizabeth L. Hartland Hayley J. Newton Nichollas E. Scott Loss of <italic toggle="yes">O</italic>-Linked Protein Glycosylation in <named-content content-type="genus-species">Burkholderia cenocepacia</named-content> Impairs Biofilm Formation and Siderophore Activity and Alters Transcriptional Regulators
description	ABSTRACT O-linked protein glycosylation is a conserved feature of the Burkholderia genus. The addition of the trisaccharide β-Gal-(1,3)-α-GalNAc-(1,3)-β-GalNAc to membrane exported proteins in Burkholderia cenocepacia is required for bacterial fitness and resistance to environmental stress. However, the underlying causes of the defects observed in the absence of glycosylation are unclear. Using proteomics, luciferase reporter assays, and DNA cross-linking, we demonstrate the loss of glycosylation leads to changes in transcriptional regulation of multiple proteins, including the repression of the master quorum CepR/I. These proteomic and transcriptional alterations lead to the abolition of biofilm formation and defects in siderophore activity. Surprisingly, the abundance of most of the known glycosylated proteins did not significantly change in the glycosylation-defective mutants, except for BCAL1086 and BCAL2974, which were found in reduced amounts, suggesting they could be degraded. However, the loss of these two proteins was not responsible for driving the proteomic alterations, biofilm formation, or siderophore activity. Together, our results show that loss of glycosylation in B. cenocepacia results in a global cell reprogramming via alteration of the transcriptional regulatory systems, which cannot be explained by the abundance changes in known B. cenocepacia glycoproteins. IMPORTANCE Protein glycosylation is increasingly recognized as a common posttranslational protein modification in bacterial species. Despite this commonality, our understanding of the role of most glycosylation systems in bacterial physiology and pathogenesis is incomplete. In this work, we investigated the effect of the disruption of O-linked glycosylation in the opportunistic pathogen Burkholderia cenocepacia using a combination of proteomic, molecular, and phenotypic assays. We find that in contrast to recent findings on the N-linked glycosylation systems of Campylobacter jejuni, O-linked glycosylation does not appear to play a role in proteome stabilization of most glycoproteins. Our results reveal that loss of glycosylation in B. cenocepacia strains leads to global proteome and transcriptional changes, including the repression of the quorum-sensing regulator cepR (BCAM1868) gene. These alterations lead to dramatic phenotypic changes in glycosylation-null strains, which are paralleled by both global proteomic and transcriptional alterations, which do not appear to directly result from the loss of glycosylation per se. This research unravels the pleiotropic effects of O-linked glycosylation in B. cenocepacia, demonstrating that its loss does not simply affect the stability of the glycoproteome, but also interferes with transcription and the broader proteome.
format	article
author	Cameron C. Oppy Leila Jebeli Miku Kuba Clare V. Oates Richard Strugnell Laura E. Edgington-Mitchell Miguel A. Valvano Elizabeth L. Hartland Hayley J. Newton Nichollas E. Scott
author_facet	Cameron C. Oppy Leila Jebeli Miku Kuba Clare V. Oates Richard Strugnell Laura E. Edgington-Mitchell Miguel A. Valvano Elizabeth L. Hartland Hayley J. Newton Nichollas E. Scott
author_sort	Cameron C. Oppy
title	Loss of <italic toggle="yes">O</italic>-Linked Protein Glycosylation in <named-content content-type="genus-species">Burkholderia cenocepacia</named-content> Impairs Biofilm Formation and Siderophore Activity and Alters Transcriptional Regulators
title_short	Loss of <italic toggle="yes">O</italic>-Linked Protein Glycosylation in <named-content content-type="genus-species">Burkholderia cenocepacia</named-content> Impairs Biofilm Formation and Siderophore Activity and Alters Transcriptional Regulators
title_full	Loss of <italic toggle="yes">O</italic>-Linked Protein Glycosylation in <named-content content-type="genus-species">Burkholderia cenocepacia</named-content> Impairs Biofilm Formation and Siderophore Activity and Alters Transcriptional Regulators
title_fullStr	Loss of <italic toggle="yes">O</italic>-Linked Protein Glycosylation in <named-content content-type="genus-species">Burkholderia cenocepacia</named-content> Impairs Biofilm Formation and Siderophore Activity and Alters Transcriptional Regulators
title_full_unstemmed	Loss of <italic toggle="yes">O</italic>-Linked Protein Glycosylation in <named-content content-type="genus-species">Burkholderia cenocepacia</named-content> Impairs Biofilm Formation and Siderophore Activity and Alters Transcriptional Regulators
title_sort	loss of <italic toggle="yes">o</italic>-linked protein glycosylation in <named-content content-type="genus-species">burkholderia cenocepacia</named-content> impairs biofilm formation and siderophore activity and alters transcriptional regulators
publisher	American Society for Microbiology
publishDate	2019
url	https://doaj.org/article/420dcd1484ec4328b4745ae031fe35ca
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Loss of <italic toggle="yes">O</italic>-Linked Protein Glycosylation in <named-content content-type="genus-species">Burkholderia cenocepacia</named-content> Impairs Biofilm Formation and Siderophore Activity and Alters Transcriptional Regulators

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