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

ABSTRACT 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 cytoplasmi...

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Autores principales:	Nadja Heinz Ernst, Analise Z. Reeves, Julia E. Ramseyer, Cammie F. Lesser
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2018
Materias:	Shigella chaperone sorting platform type III secretion system Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/9676791c5a064467976443427a1194e0
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spelling	oai:doaj.org-article:9676791c5a064467976443427a1194e02021-11-15T16:00:24ZHigh-Throughput Screening of Type III Secretion Determinants Reveals a Major Chaperone-Independent Pathway10.1128/mBio.01050-182150-7511https://doaj.org/article/9676791c5a064467976443427a1194e02018-07-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.01050-18https://doaj.org/toc/2150-7511ABSTRACT 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. IMPORTANCE Many bacterial pathogens use specialized nanomachines, including type III secretion systems, to directly inject virulence proteins (effectors) into host cells. Here, we present the first extensive analysis of chaperone dependence in the process of type III effector secretion, providing strong evidence for the existence of a previously unrecognized chaperone-independent pathway. This noncanonical pathway is likely common to many bacteria, as an extensive review of the literature reveals that the secretion of multiple type III effectors has not yet been knowingly linked to a chaperone. While additional studies will be required to discern the molecular details of this pathway, its prevalence suggests that it can likely serve as a new target for the development of antimicrobial agents.Nadja Heinz ErnstAnalise Z. ReevesJulia E. RamseyerCammie F. LesserAmerican Society for MicrobiologyarticleShigellachaperonesorting platformtype III secretion systemMicrobiologyQR1-502ENmBio, Vol 9, Iss 3 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	Shigella chaperone sorting platform type III secretion system Microbiology QR1-502
spellingShingle	Shigella chaperone sorting platform type III secretion system Microbiology QR1-502 Nadja Heinz Ernst Analise Z. Reeves Julia E. Ramseyer Cammie F. Lesser High-Throughput Screening of Type III Secretion Determinants Reveals a Major Chaperone-Independent Pathway
description	ABSTRACT 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. IMPORTANCE Many bacterial pathogens use specialized nanomachines, including type III secretion systems, to directly inject virulence proteins (effectors) into host cells. Here, we present the first extensive analysis of chaperone dependence in the process of type III effector secretion, providing strong evidence for the existence of a previously unrecognized chaperone-independent pathway. This noncanonical pathway is likely common to many bacteria, as an extensive review of the literature reveals that the secretion of multiple type III effectors has not yet been knowingly linked to a chaperone. While additional studies will be required to discern the molecular details of this pathway, its prevalence suggests that it can likely serve as a new target for the development of antimicrobial agents.
format	article
author	Nadja Heinz Ernst Analise Z. Reeves Julia E. Ramseyer Cammie F. Lesser
author_facet	Nadja Heinz Ernst Analise Z. Reeves Julia E. Ramseyer Cammie F. Lesser
author_sort	Nadja Heinz Ernst
title	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_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_sort	high-throughput screening of type iii secretion determinants reveals a major chaperone-independent pathway
publisher	American Society for Microbiology
publishDate	2018
url	https://doaj.org/article/9676791c5a064467976443427a1194e0
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High-Throughput Screening of Type III Secretion Determinants Reveals a Major Chaperone-Independent Pathway

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