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Aberrant Membrane Structures in Hypervesiculating Escherichia coli Strain ΔmlaEΔnlpI Visualized by Electron Microscopy

Escherichia coli produces extracellular vesicles called outer membrane vesicles (OMVs) by releasing a part of its outer membrane. We previously reported that the combined deletion of nlpI and mlaE, related to envelope structure and phospholipid accumulation in the outer leaflet of the outer membrane...

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Autores principales: Ojima, Yoshihiro, Sawabe, Tomomi, Nakagawa, Mao, Tahara, Yuhei O., Miyata, Makoto, Azuma, Masayuki
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8386018/
https://www.ncbi.nlm.nih.gov/pubmed/34456889
http://dx.doi.org/10.3389/fmicb.2021.706525
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author Ojima, Yoshihiro
Sawabe, Tomomi
Nakagawa, Mao
Tahara, Yuhei O.
Miyata, Makoto
Azuma, Masayuki
author_facet Ojima, Yoshihiro
Sawabe, Tomomi
Nakagawa, Mao
Tahara, Yuhei O.
Miyata, Makoto
Azuma, Masayuki
author_sort Ojima, Yoshihiro
collection PubMed
description Escherichia coli produces extracellular vesicles called outer membrane vesicles (OMVs) by releasing a part of its outer membrane. We previously reported that the combined deletion of nlpI and mlaE, related to envelope structure and phospholipid accumulation in the outer leaflet of the outer membrane, respectively, resulted in the synergistic increase of OMV production. In this study, the analysis of ΔmlaEΔnlpI cells using quick-freeze, deep-etch electron microscopy (QFDE-EM) revealed that plasmolysis occurred at the tip of the long axis in cells and that OMVs formed from this tip. Plasmolysis was also observed in the single-gene knockout mutants ΔnlpI and ΔmlaE. This study has demonstrated that plasmolysis was induced in the hypervesiculating mutant E. coli cells. Furthermore, intracellular vesicles and multilamellar OMV were observed in the ΔmlaEΔnlpI cells. Meanwhile, the secretion of recombinant green fluorescent protein (GFP) expressed in the cytosol of the ΔmlaEΔnlpI cells was more than 100 times higher than that of WT and ΔnlpI, and about 50 times higher than that of ΔmlaE in the OMV fraction, suggesting that cytosolic components were incorporated into outer-inner membrane vesicles (OIMVs) and released into the extracellular space. Additionally, QFDE-EM analysis revealed that ΔmlaEΔnlpI sacculi contained many holes noticeably larger than the mean radius of the peptidoglycan (PG) pores in wild-type (WT) E. coli. These results suggest that in ΔmlaEΔnlpI cells, cytoplasmic membrane materials protrude into the periplasmic space through the peptidoglycan holes and are released as OIMVs.
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spelling pubmed-83860182021-08-26 Aberrant Membrane Structures in Hypervesiculating Escherichia coli Strain ΔmlaEΔnlpI Visualized by Electron Microscopy Ojima, Yoshihiro Sawabe, Tomomi Nakagawa, Mao Tahara, Yuhei O. Miyata, Makoto Azuma, Masayuki Front Microbiol Microbiology Escherichia coli produces extracellular vesicles called outer membrane vesicles (OMVs) by releasing a part of its outer membrane. We previously reported that the combined deletion of nlpI and mlaE, related to envelope structure and phospholipid accumulation in the outer leaflet of the outer membrane, respectively, resulted in the synergistic increase of OMV production. In this study, the analysis of ΔmlaEΔnlpI cells using quick-freeze, deep-etch electron microscopy (QFDE-EM) revealed that plasmolysis occurred at the tip of the long axis in cells and that OMVs formed from this tip. Plasmolysis was also observed in the single-gene knockout mutants ΔnlpI and ΔmlaE. This study has demonstrated that plasmolysis was induced in the hypervesiculating mutant E. coli cells. Furthermore, intracellular vesicles and multilamellar OMV were observed in the ΔmlaEΔnlpI cells. Meanwhile, the secretion of recombinant green fluorescent protein (GFP) expressed in the cytosol of the ΔmlaEΔnlpI cells was more than 100 times higher than that of WT and ΔnlpI, and about 50 times higher than that of ΔmlaE in the OMV fraction, suggesting that cytosolic components were incorporated into outer-inner membrane vesicles (OIMVs) and released into the extracellular space. Additionally, QFDE-EM analysis revealed that ΔmlaEΔnlpI sacculi contained many holes noticeably larger than the mean radius of the peptidoglycan (PG) pores in wild-type (WT) E. coli. These results suggest that in ΔmlaEΔnlpI cells, cytoplasmic membrane materials protrude into the periplasmic space through the peptidoglycan holes and are released as OIMVs. Frontiers Media S.A. 2021-08-11 /pmc/articles/PMC8386018/ /pubmed/34456889 http://dx.doi.org/10.3389/fmicb.2021.706525 Text en Copyright © 2021 Ojima, Sawabe, Nakagawa, Tahara, Miyata and Azuma. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Microbiology
Ojima, Yoshihiro
Sawabe, Tomomi
Nakagawa, Mao
Tahara, Yuhei O.
Miyata, Makoto
Azuma, Masayuki
Aberrant Membrane Structures in Hypervesiculating Escherichia coli Strain ΔmlaEΔnlpI Visualized by Electron Microscopy
title Aberrant Membrane Structures in Hypervesiculating Escherichia coli Strain ΔmlaEΔnlpI Visualized by Electron Microscopy
title_full Aberrant Membrane Structures in Hypervesiculating Escherichia coli Strain ΔmlaEΔnlpI Visualized by Electron Microscopy
title_fullStr Aberrant Membrane Structures in Hypervesiculating Escherichia coli Strain ΔmlaEΔnlpI Visualized by Electron Microscopy
title_full_unstemmed Aberrant Membrane Structures in Hypervesiculating Escherichia coli Strain ΔmlaEΔnlpI Visualized by Electron Microscopy
title_short Aberrant Membrane Structures in Hypervesiculating Escherichia coli Strain ΔmlaEΔnlpI Visualized by Electron Microscopy
title_sort aberrant membrane structures in hypervesiculating escherichia coli strain δmlaeδnlpi visualized by electron microscopy
topic Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8386018/
https://www.ncbi.nlm.nih.gov/pubmed/34456889
http://dx.doi.org/10.3389/fmicb.2021.706525
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