Outer membrane vesicles from the probiotic Escherichia coli Nissle 1917 and the commensal ECOR12 enter intestinal epithelial cells via clathrin-dependent endocytosis and elicit differential effects on DNA damage.

Interactions between intestinal microbiota and the human host are complex. The gut mucosal surface is covered by a mucin layer that prevents bacteria from accessing the epithelial cells. Thus, the crosstalk between microbiota and the host mainly rely on secreted factors that can go through the mucus...

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Autores: Cañas, María Alexandra, Giménez Claudio, Rosa, Fábrega Fernández, María José, Toloza Maturana, Lorena, Baldomà Llavinés, Laura, Badía Palacín, Josefa
Tipo de recurso: artículo
Estado:Versión publicada
Fecha de publicación:2016
País:España
Institución:Universidad de Barcelona
Repositorio:Dipòsit Digital de la UB
OAI Identifier:oai:diposit.ub.edu:2445/104299
Acceso en línea:https://hdl.handle.net/2445/104299
Access Level:acceso abierto
Palabra clave:Escheríchia coli
Probiòtics
Intestins
Escherichia coli
Probiotics
Intestines
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spelling Outer membrane vesicles from the probiotic Escherichia coli Nissle 1917 and the commensal ECOR12 enter intestinal epithelial cells via clathrin-dependent endocytosis and elicit differential effects on DNA damage.Cañas, María AlexandraGiménez Claudio, RosaFábrega Fernández, María JoséToloza Maturana, LorenaBaldomà Llavinés, LauraBadía Palacín, JosefaEscheríchia coliProbiòticsIntestinsEscherichia coliProbioticsIntestinesInteractions between intestinal microbiota and the human host are complex. The gut mucosal surface is covered by a mucin layer that prevents bacteria from accessing the epithelial cells. Thus, the crosstalk between microbiota and the host mainly rely on secreted factors that can go through the mucus layer and reach the epithelium. In this context, vesicles released by commensal strains are seen as key players in signaling processes in the intestinal mucosa. Studies with Gram-negative pathogens showed that outer membrane vesicles (OMVs) are internalized into the host cell by endocytosis, but the entry mechanism for microbiota-derived vesicles is unknown. Escherichia coli strains are found as part of normal human gut microbiota. In this work, we elucidate the pathway that mediate internalization of OMVs from the probiotic E.coli Nissle 1917 (EcN) and the commensal ECOR12 strains in several human intestinal epithelial cell lines. Time course measurement of fluorescence and microscopy analysis performed with rhodamine B-R18-labeled OMVs in the presence of endocytosis inhibitors showed that OMVs from these strains enter epithelial cells via clathrin-mediated endocytosis. Vesicles use the same endocytosis pathway in polarized epithelial monolayers. Internalized OMVs are sorted to lysosomal compartments as shown by their colocalization with clathrin and specific markers of endosomes and lysosomes. OMVs from both strains did not affect cell viability, but reduce proliferation of HT-29 cells. Labeling of 8-oxo-dG adducts in DNA revealed that neither OMVs from EcN nor from ECOR12 promoted oxidative DNA damage. In contrast, flow cytometry analysis of phosphorylated γH2AX evidenced that OMVs from the probiotic EcN significantly produced more double strand breaks in DNA than ECOR12 OMVs. The EcN genotoxic effects have been attributed to the synthesis of colibactin. However, it is not known how colibactin is exported and delivered into host cells. Whether colibactin is secreted via OMVs is an open question that needs further study.Public Library of Science (PLoS)2016info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionapplication/pdfhttps://hdl.handle.net/2445/104299Articles publicats en revistes (Bioquímica i Fisiologia)reponame:Dipòsit Digital de la UBinstname:Universidad de BarcelonaInglésReproducció del document publicat a: https://doi.org/10.1371/journal.pone.0160374PLoS One, 2016, vol. 11, num. 8, p. e0160374https://doi.org/10.1371/journal.pone.0160374cc-by (c) Cañas Pacheco, Maria Alexandra et al., 2016http://creativecommons.org/licenses/by/3.0/esinfo:eu-repo/semantics/openAccessoai:diposit.ub.edu:2445/1042992026-05-27T06:46:51Z
dc.title.none.fl_str_mv Outer membrane vesicles from the probiotic Escherichia coli Nissle 1917 and the commensal ECOR12 enter intestinal epithelial cells via clathrin-dependent endocytosis and elicit differential effects on DNA damage.
title Outer membrane vesicles from the probiotic Escherichia coli Nissle 1917 and the commensal ECOR12 enter intestinal epithelial cells via clathrin-dependent endocytosis and elicit differential effects on DNA damage.
spellingShingle Outer membrane vesicles from the probiotic Escherichia coli Nissle 1917 and the commensal ECOR12 enter intestinal epithelial cells via clathrin-dependent endocytosis and elicit differential effects on DNA damage.
Cañas, María Alexandra
Escheríchia coli
Probiòtics
Intestins
Escherichia coli
Probiotics
Intestines
title_short Outer membrane vesicles from the probiotic Escherichia coli Nissle 1917 and the commensal ECOR12 enter intestinal epithelial cells via clathrin-dependent endocytosis and elicit differential effects on DNA damage.
title_full Outer membrane vesicles from the probiotic Escherichia coli Nissle 1917 and the commensal ECOR12 enter intestinal epithelial cells via clathrin-dependent endocytosis and elicit differential effects on DNA damage.
title_fullStr Outer membrane vesicles from the probiotic Escherichia coli Nissle 1917 and the commensal ECOR12 enter intestinal epithelial cells via clathrin-dependent endocytosis and elicit differential effects on DNA damage.
title_full_unstemmed Outer membrane vesicles from the probiotic Escherichia coli Nissle 1917 and the commensal ECOR12 enter intestinal epithelial cells via clathrin-dependent endocytosis and elicit differential effects on DNA damage.
title_sort Outer membrane vesicles from the probiotic Escherichia coli Nissle 1917 and the commensal ECOR12 enter intestinal epithelial cells via clathrin-dependent endocytosis and elicit differential effects on DNA damage.
dc.creator.none.fl_str_mv Cañas, María Alexandra
Giménez Claudio, Rosa
Fábrega Fernández, María José
Toloza Maturana, Lorena
Baldomà Llavinés, Laura
Badía Palacín, Josefa
author Cañas, María Alexandra
author_facet Cañas, María Alexandra
Giménez Claudio, Rosa
Fábrega Fernández, María José
Toloza Maturana, Lorena
Baldomà Llavinés, Laura
Badía Palacín, Josefa
author_role author
author2 Giménez Claudio, Rosa
Fábrega Fernández, María José
Toloza Maturana, Lorena
Baldomà Llavinés, Laura
Badía Palacín, Josefa
author2_role author
author
author
author
author
dc.subject.none.fl_str_mv Escheríchia coli
Probiòtics
Intestins
Escherichia coli
Probiotics
Intestines
topic Escheríchia coli
Probiòtics
Intestins
Escherichia coli
Probiotics
Intestines
description Interactions between intestinal microbiota and the human host are complex. The gut mucosal surface is covered by a mucin layer that prevents bacteria from accessing the epithelial cells. Thus, the crosstalk between microbiota and the host mainly rely on secreted factors that can go through the mucus layer and reach the epithelium. In this context, vesicles released by commensal strains are seen as key players in signaling processes in the intestinal mucosa. Studies with Gram-negative pathogens showed that outer membrane vesicles (OMVs) are internalized into the host cell by endocytosis, but the entry mechanism for microbiota-derived vesicles is unknown. Escherichia coli strains are found as part of normal human gut microbiota. In this work, we elucidate the pathway that mediate internalization of OMVs from the probiotic E.coli Nissle 1917 (EcN) and the commensal ECOR12 strains in several human intestinal epithelial cell lines. Time course measurement of fluorescence and microscopy analysis performed with rhodamine B-R18-labeled OMVs in the presence of endocytosis inhibitors showed that OMVs from these strains enter epithelial cells via clathrin-mediated endocytosis. Vesicles use the same endocytosis pathway in polarized epithelial monolayers. Internalized OMVs are sorted to lysosomal compartments as shown by their colocalization with clathrin and specific markers of endosomes and lysosomes. OMVs from both strains did not affect cell viability, but reduce proliferation of HT-29 cells. Labeling of 8-oxo-dG adducts in DNA revealed that neither OMVs from EcN nor from ECOR12 promoted oxidative DNA damage. In contrast, flow cytometry analysis of phosphorylated γH2AX evidenced that OMVs from the probiotic EcN significantly produced more double strand breaks in DNA than ECOR12 OMVs. The EcN genotoxic effects have been attributed to the synthesis of colibactin. However, it is not known how colibactin is exported and delivered into host cells. Whether colibactin is secreted via OMVs is an open question that needs further study.
publishDate 2016
dc.date.none.fl_str_mv 2016
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv https://hdl.handle.net/2445/104299
url https://hdl.handle.net/2445/104299
dc.language.none.fl_str_mv Inglés
language_invalid_str_mv Inglés
dc.relation.none.fl_str_mv Reproducció del document publicat a: https://doi.org/10.1371/journal.pone.0160374
PLoS One, 2016, vol. 11, num. 8, p. e0160374
https://doi.org/10.1371/journal.pone.0160374
dc.rights.none.fl_str_mv cc-by (c) Cañas Pacheco, Maria Alexandra et al., 2016
http://creativecommons.org/licenses/by/3.0/es
info:eu-repo/semantics/openAccess
rights_invalid_str_mv cc-by (c) Cañas Pacheco, Maria Alexandra et al., 2016
http://creativecommons.org/licenses/by/3.0/es
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.publisher.none.fl_str_mv Public Library of Science (PLoS)
publisher.none.fl_str_mv Public Library of Science (PLoS)
dc.source.none.fl_str_mv Articles publicats en revistes (Bioquímica i Fisiologia)
reponame:Dipòsit Digital de la UB
instname:Universidad de Barcelona
instname_str Universidad de Barcelona
reponame_str Dipòsit Digital de la UB
collection Dipòsit Digital de la UB
repository.name.fl_str_mv
repository.mail.fl_str_mv
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score 15,298079

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