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High-Throughput Screening of Type III Secretion Determinants Reveals a Major Chaperone-Independent Pathway

Numerous Gram-negative bacterial pathogens utilize type III secretion systems (T3SSs) to inject tens of effector proteins directly into the cytosol of host cells. Through interactions with cognate chaperones, type III effectors are defined and recruited to the sorting platform, a cytoplasmic compone...

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Detalles Bibliográficos
Autores principales: Ernst, Nadja Heinz, Reeves, Analise Z., Ramseyer, Julia E., Lesser, Cammie F.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Society for Microbiology 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6016238/
https://www.ncbi.nlm.nih.gov/pubmed/29921672
http://dx.doi.org/10.1128/mBio.01050-18
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author Ernst, Nadja Heinz
Reeves, Analise Z.
Ramseyer, Julia E.
Lesser, Cammie F.
author_facet Ernst, Nadja Heinz
Reeves, Analise Z.
Ramseyer, Julia E.
Lesser, Cammie F.
author_sort Ernst, Nadja Heinz
collection PubMed
description Numerous Gram-negative bacterial pathogens utilize type III secretion systems (T3SSs) to inject tens of effector proteins directly into the cytosol of host cells. Through interactions with cognate chaperones, type III effectors are defined and recruited to the sorting platform, a cytoplasmic component of these membrane-embedded nanomachines. However, notably, a comprehensive review of the literature reveals that the secretion of most type III effectors has not yet been linked to a chaperone, raising questions regarding the existence of unknown chaperones as well as the universality of chaperones in effector secretion. Here, we describe the development of the first high-throughput type III secretion (T3S) assay, a semiautomated solid-plate-based assay, which enables the side-by-side comparison of secretion of over 20 Shigella effectors under a multitude of conditions. Strikingly, we found that the majority of Shigella effectors are secreted at equivalent levels by wild-type and variants of Shigella that no longer encode one or all known Shigella T3S effector chaperones. In addition, we found that Shigella effectors are efficiently secreted from a laboratory strain of Escherichia coli expressing the core Shigella type III secretion apparatus (T3SA) but no other Shigella-specific proteins. Furthermore, we observed that the sequences necessary and sufficient to define chaperone-dependent and -independent effectors are fundamentally different. Together, these findings support the existence of a major, previously unrecognized, noncanonical chaperone-independent secretion pathway that is likely common to many T3SSs.
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spelling pubmed-60162382018-06-26 High-Throughput Screening of Type III Secretion Determinants Reveals a Major Chaperone-Independent Pathway Ernst, Nadja Heinz Reeves, Analise Z. Ramseyer, Julia E. Lesser, Cammie F. mBio Research Article Numerous Gram-negative bacterial pathogens utilize type III secretion systems (T3SSs) to inject tens of effector proteins directly into the cytosol of host cells. Through interactions with cognate chaperones, type III effectors are defined and recruited to the sorting platform, a cytoplasmic component of these membrane-embedded nanomachines. However, notably, a comprehensive review of the literature reveals that the secretion of most type III effectors has not yet been linked to a chaperone, raising questions regarding the existence of unknown chaperones as well as the universality of chaperones in effector secretion. Here, we describe the development of the first high-throughput type III secretion (T3S) assay, a semiautomated solid-plate-based assay, which enables the side-by-side comparison of secretion of over 20 Shigella effectors under a multitude of conditions. Strikingly, we found that the majority of Shigella effectors are secreted at equivalent levels by wild-type and variants of Shigella that no longer encode one or all known Shigella T3S effector chaperones. In addition, we found that Shigella effectors are efficiently secreted from a laboratory strain of Escherichia coli expressing the core Shigella type III secretion apparatus (T3SA) but no other Shigella-specific proteins. Furthermore, we observed that the sequences necessary and sufficient to define chaperone-dependent and -independent effectors are fundamentally different. Together, these findings support the existence of a major, previously unrecognized, noncanonical chaperone-independent secretion pathway that is likely common to many T3SSs. American Society for Microbiology 2018-06-19 /pmc/articles/PMC6016238/ /pubmed/29921672 http://dx.doi.org/10.1128/mBio.01050-18 Text en Copyright © 2018 Ernst et al. https://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license (https://creativecommons.org/licenses/by/4.0/) .
spellingShingle Research Article
Ernst, Nadja Heinz
Reeves, Analise Z.
Ramseyer, Julia E.
Lesser, Cammie F.
High-Throughput Screening of Type III Secretion Determinants Reveals a Major Chaperone-Independent Pathway
title High-Throughput Screening of Type III Secretion Determinants Reveals a Major Chaperone-Independent Pathway
title_full High-Throughput Screening of Type III Secretion Determinants Reveals a Major Chaperone-Independent Pathway
title_fullStr High-Throughput Screening of Type III Secretion Determinants Reveals a Major Chaperone-Independent Pathway
title_full_unstemmed High-Throughput Screening of Type III Secretion Determinants Reveals a Major Chaperone-Independent Pathway
title_short High-Throughput Screening of Type III Secretion Determinants Reveals a Major Chaperone-Independent Pathway
title_sort high-throughput screening of type iii secretion determinants reveals a major chaperone-independent pathway
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6016238/
https://www.ncbi.nlm.nih.gov/pubmed/29921672
http://dx.doi.org/10.1128/mBio.01050-18
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