Incorporation of Plasmid DNA Into Bacterial Membrane Vesicles by Peptidoglycan Defects in Escherichia coli

Membrane vesicles (MVs) are released by various prokaryotes and play a role in the delivery of various cell-cell interaction factors. Recent studies have determined that these vesicles are capable of functioning as mediators of horizontal gene transfer. Outer membrane vesicles (OMVs) are a type of M...

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Autores principales:	Sharmin Aktar, Yuhi Okamoto, So Ueno, Yuhei O. Tahara, Masayoshi Imaizumi, Masaki Shintani, Makoto Miyata, Hiroyuki Futamata, Hideaki Nojiri, Yosuke Tashiro
Formato:	article
Lenguaje:	EN
Publicado:	Frontiers Media S.A. 2021
Materias:	membrane vesicles plasmid peptidoglycan glycine quick-freeze deep-etch and replica electron microscopy outer membrane vesicles Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/6538497dd6444f93a96345e304de6b6e
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spelling	oai:doaj.org-article:6538497dd6444f93a96345e304de6b6e2021-12-01T13:53:57ZIncorporation of Plasmid DNA Into Bacterial Membrane Vesicles by Peptidoglycan Defects in Escherichia coli1664-302X10.3389/fmicb.2021.747606https://doaj.org/article/6538497dd6444f93a96345e304de6b6e2021-11-01T00:00:00Zhttps://www.frontiersin.org/articles/10.3389/fmicb.2021.747606/fullhttps://doaj.org/toc/1664-302XMembrane vesicles (MVs) are released by various prokaryotes and play a role in the delivery of various cell-cell interaction factors. Recent studies have determined that these vesicles are capable of functioning as mediators of horizontal gene transfer. Outer membrane vesicles (OMVs) are a type of MV that is released by Gram-negative bacteria and primarily composed of outer membrane and periplasm components; however, it remains largely unknown why DNA is contained within OMVs. Our study aimed to understand the mechanism by which DNA that is localized in the cytoplasm is incorporated into OMVs in Gram-negative bacteria. We compared DNA associated with OMVs using Escherichia coli BW25113 cells harboring the non-conjugative, non-mobilized, and high-copy plasmid pUC19 and its hypervesiculating mutants that included ΔnlpI, ΔrseA, and ΔtolA. Plasmid copy per vesicle was increased in OMVs derived from ΔnlpI, in which peptidoglycan (PG) breakdown and synthesis are altered. When supplemented with 1% glycine to inhibit PG synthesis, both OMV formation and plasmid copy per vesicle were increased in the wild type. The bacterial membrane condition test indicated that membrane permeability was increased in the presence of glycine at the late exponential phase, in which cell lysis did not occur. Additionally, quick-freeze deep-etch and replica electron microscopy observations revealed that outer-inner membrane vesicles (O-IMVs) are formed in the presence of glycine. Thus, two proposed routes for DNA incorporation into OMVs under PG-damaged conditions are suggested. These routes include DNA leakage due to increased membrane permeation and O-IMV formation. Additionally, our findings contribute to a greater understanding of the vesicle-mediated horizontal gene transfer that occurs in nature and the utilization of MVs for DNA cargo.Sharmin AktarYuhi OkamotoSo UenoYuhei O. TaharaYuhei O. TaharaMasayoshi ImaizumiMasaki ShintaniMasaki ShintaniMasaki ShintaniMasaki ShintaniMakoto MiyataMakoto MiyataHiroyuki FutamataHiroyuki FutamataHiroyuki FutamataHiroyuki FutamataHideaki NojiriYosuke TashiroYosuke TashiroYosuke TashiroYosuke TashiroFrontiers Media S.A.articlemembrane vesiclesplasmidpeptidoglycanglycinequick-freeze deep-etch and replica electron microscopyouter membrane vesiclesMicrobiologyQR1-502ENFrontiers in Microbiology, Vol 12 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	membrane vesicles plasmid peptidoglycan glycine quick-freeze deep-etch and replica electron microscopy outer membrane vesicles Microbiology QR1-502
spellingShingle	membrane vesicles plasmid peptidoglycan glycine quick-freeze deep-etch and replica electron microscopy outer membrane vesicles Microbiology QR1-502 Sharmin Aktar Yuhi Okamoto So Ueno Yuhei O. Tahara Yuhei O. Tahara Masayoshi Imaizumi Masaki Shintani Masaki Shintani Masaki Shintani Masaki Shintani Makoto Miyata Makoto Miyata Hiroyuki Futamata Hiroyuki Futamata Hiroyuki Futamata Hiroyuki Futamata Hideaki Nojiri Yosuke Tashiro Yosuke Tashiro Yosuke Tashiro Yosuke Tashiro Incorporation of Plasmid DNA Into Bacterial Membrane Vesicles by Peptidoglycan Defects in Escherichia coli
description	Membrane vesicles (MVs) are released by various prokaryotes and play a role in the delivery of various cell-cell interaction factors. Recent studies have determined that these vesicles are capable of functioning as mediators of horizontal gene transfer. Outer membrane vesicles (OMVs) are a type of MV that is released by Gram-negative bacteria and primarily composed of outer membrane and periplasm components; however, it remains largely unknown why DNA is contained within OMVs. Our study aimed to understand the mechanism by which DNA that is localized in the cytoplasm is incorporated into OMVs in Gram-negative bacteria. We compared DNA associated with OMVs using Escherichia coli BW25113 cells harboring the non-conjugative, non-mobilized, and high-copy plasmid pUC19 and its hypervesiculating mutants that included ΔnlpI, ΔrseA, and ΔtolA. Plasmid copy per vesicle was increased in OMVs derived from ΔnlpI, in which peptidoglycan (PG) breakdown and synthesis are altered. When supplemented with 1% glycine to inhibit PG synthesis, both OMV formation and plasmid copy per vesicle were increased in the wild type. The bacterial membrane condition test indicated that membrane permeability was increased in the presence of glycine at the late exponential phase, in which cell lysis did not occur. Additionally, quick-freeze deep-etch and replica electron microscopy observations revealed that outer-inner membrane vesicles (O-IMVs) are formed in the presence of glycine. Thus, two proposed routes for DNA incorporation into OMVs under PG-damaged conditions are suggested. These routes include DNA leakage due to increased membrane permeation and O-IMV formation. Additionally, our findings contribute to a greater understanding of the vesicle-mediated horizontal gene transfer that occurs in nature and the utilization of MVs for DNA cargo.
format	article
author	Sharmin Aktar Yuhi Okamoto So Ueno Yuhei O. Tahara Yuhei O. Tahara Masayoshi Imaizumi Masaki Shintani Masaki Shintani Masaki Shintani Masaki Shintani Makoto Miyata Makoto Miyata Hiroyuki Futamata Hiroyuki Futamata Hiroyuki Futamata Hiroyuki Futamata Hideaki Nojiri Yosuke Tashiro Yosuke Tashiro Yosuke Tashiro Yosuke Tashiro
author_facet	Sharmin Aktar Yuhi Okamoto So Ueno Yuhei O. Tahara Yuhei O. Tahara Masayoshi Imaizumi Masaki Shintani Masaki Shintani Masaki Shintani Masaki Shintani Makoto Miyata Makoto Miyata Hiroyuki Futamata Hiroyuki Futamata Hiroyuki Futamata Hiroyuki Futamata Hideaki Nojiri Yosuke Tashiro Yosuke Tashiro Yosuke Tashiro Yosuke Tashiro
author_sort	Sharmin Aktar
title	Incorporation of Plasmid DNA Into Bacterial Membrane Vesicles by Peptidoglycan Defects in Escherichia coli
title_short	Incorporation of Plasmid DNA Into Bacterial Membrane Vesicles by Peptidoglycan Defects in Escherichia coli
title_full	Incorporation of Plasmid DNA Into Bacterial Membrane Vesicles by Peptidoglycan Defects in Escherichia coli
title_fullStr	Incorporation of Plasmid DNA Into Bacterial Membrane Vesicles by Peptidoglycan Defects in Escherichia coli
title_full_unstemmed	Incorporation of Plasmid DNA Into Bacterial Membrane Vesicles by Peptidoglycan Defects in Escherichia coli
title_sort	incorporation of plasmid dna into bacterial membrane vesicles by peptidoglycan defects in escherichia coli
publisher	Frontiers Media S.A.
publishDate	2021
url	https://doaj.org/article/6538497dd6444f93a96345e304de6b6e
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Incorporation of Plasmid DNA Into Bacterial Membrane Vesicles by Peptidoglycan Defects in Escherichia coli

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