Virulence of the Pathogen <named-content content-type="genus-species">Porphyromonas gingivalis</named-content> Is Controlled by the CRISPR-Cas Protein Cas3

ABSTRACT The CRISPR (clustered regularly interspaced short palindromic repeat)-Cas system is a unique genomic entity that provides prokaryotic cells with adaptive and heritable immunity. Initial studies identified CRISPRs as central elements used by bacteria to protect against foreign nucleic acids;...

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Autores principales:	Jose Solbiati, Ana Duran-Pinedo, Fernanda Godoy Rocha, Frank C. Gibson, Jorge Frias-Lopez
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2020
Materias:	CRISPR-Cas virulence pathogenesis periodontal disease Porphyromonas gingivalis Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/7bfecfb23ba64cbfa3d4bda9e35cc9bb
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spelling	oai:doaj.org-article:7bfecfb23ba64cbfa3d4bda9e35cc9bb2021-12-02T18:44:36ZVirulence of the Pathogen <named-content content-type="genus-species">Porphyromonas gingivalis</named-content> Is Controlled by the CRISPR-Cas Protein Cas310.1128/mSystems.00852-202379-5077https://doaj.org/article/7bfecfb23ba64cbfa3d4bda9e35cc9bb2020-10-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mSystems.00852-20https://doaj.org/toc/2379-5077ABSTRACT The CRISPR (clustered regularly interspaced short palindromic repeat)-Cas system is a unique genomic entity that provides prokaryotic cells with adaptive and heritable immunity. Initial studies identified CRISPRs as central elements used by bacteria to protect against foreign nucleic acids; however, emerging evidence points to CRISPR involvement in bacterial virulence. The present study aimed to identify the participation of one CRISPR-Cas protein, Cas3, in the virulence of the oral pathogen Porphyromonas gingivalis, an organism highly associated with periodontitis. Our results show that compared to the wild type, a mutant with a deletion of the Cas3 gene, an essential nuclease part of the class 1 type I CRISPR-Cas system, increased the virulence of P. gingivalis. In vitro infection modeling revealed only mildly enhanced production of proinflammatory cytokines by THP-1 cells when infected with the mutant strain. Dual transcriptome sequencing (RNA-seq) analysis of infected THP-1 cells showed an increase in expression of genes associated with pathogenesis in response to Δcas3 mutant infection, with the target of Cas3 activities in neutrophil chemotaxis and gene silencing. The importance of cas3 in controlling virulence was corroborated in a Galleria mellonella infection model, where the presence of the Δcas3 mutant resulted in a statistically significant increase in mortality of G. mellonella. A time-series analysis of transcription patterning during infection showed that G. mellonella elicited very different immune responses to the wild-type and the Δcas3 mutant strains and revealed a rearrangement of association in coexpression networks. Together, these observations show for the first time that Cas3 plays a significant role in regulating the virulence of P. gingivalis. IMPORTANCE Porphyromonas gingivalis is a key pathogen of periodontitis, a polymicrobial disease characterized by a chronic inflammation that destroys the tissues supporting the teeth. Thus, understanding the virulence potential of P. gingivalis is essential to maintaining a healthy oral microbiome. In nonoral organisms, CRISPR-Cas systems have been shown to modulate a variety of microbial processes, including protection from exogenous nucleic acids, and, more recently, have been implicated in bacterial virulence. Previously, our clinical findings identified activation of the CRISPR-Cas system in patient samples at the transition to disease; however, the mechanism of contribution to disease remained unknown. The importance of the present study resides in that it is becoming increasingly clear that CRISPR-associated proteins have broader functions than initially thought and that those functions now include their role in the virulence of periodontal pathogens. Studying a P. gingivalis cas3 mutant, we demonstrate that at least one of the CRISPR-Cas systems is involved in the regulation of virulence during infection.Jose SolbiatiAna Duran-PinedoFernanda Godoy RochaFrank C. GibsonJorge Frias-LopezAmerican Society for MicrobiologyarticleCRISPR-Casvirulencepathogenesisperiodontal diseasePorphyromonas gingivalisMicrobiologyQR1-502ENmSystems, Vol 5, Iss 5 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	CRISPR-Cas virulence pathogenesis periodontal disease Porphyromonas gingivalis Microbiology QR1-502
spellingShingle	CRISPR-Cas virulence pathogenesis periodontal disease Porphyromonas gingivalis Microbiology QR1-502 Jose Solbiati Ana Duran-Pinedo Fernanda Godoy Rocha Frank C. Gibson Jorge Frias-Lopez Virulence of the Pathogen <named-content content-type="genus-species">Porphyromonas gingivalis</named-content> Is Controlled by the CRISPR-Cas Protein Cas3
description	ABSTRACT The CRISPR (clustered regularly interspaced short palindromic repeat)-Cas system is a unique genomic entity that provides prokaryotic cells with adaptive and heritable immunity. Initial studies identified CRISPRs as central elements used by bacteria to protect against foreign nucleic acids; however, emerging evidence points to CRISPR involvement in bacterial virulence. The present study aimed to identify the participation of one CRISPR-Cas protein, Cas3, in the virulence of the oral pathogen Porphyromonas gingivalis, an organism highly associated with periodontitis. Our results show that compared to the wild type, a mutant with a deletion of the Cas3 gene, an essential nuclease part of the class 1 type I CRISPR-Cas system, increased the virulence of P. gingivalis. In vitro infection modeling revealed only mildly enhanced production of proinflammatory cytokines by THP-1 cells when infected with the mutant strain. Dual transcriptome sequencing (RNA-seq) analysis of infected THP-1 cells showed an increase in expression of genes associated with pathogenesis in response to Δcas3 mutant infection, with the target of Cas3 activities in neutrophil chemotaxis and gene silencing. The importance of cas3 in controlling virulence was corroborated in a Galleria mellonella infection model, where the presence of the Δcas3 mutant resulted in a statistically significant increase in mortality of G. mellonella. A time-series analysis of transcription patterning during infection showed that G. mellonella elicited very different immune responses to the wild-type and the Δcas3 mutant strains and revealed a rearrangement of association in coexpression networks. Together, these observations show for the first time that Cas3 plays a significant role in regulating the virulence of P. gingivalis. IMPORTANCE Porphyromonas gingivalis is a key pathogen of periodontitis, a polymicrobial disease characterized by a chronic inflammation that destroys the tissues supporting the teeth. Thus, understanding the virulence potential of P. gingivalis is essential to maintaining a healthy oral microbiome. In nonoral organisms, CRISPR-Cas systems have been shown to modulate a variety of microbial processes, including protection from exogenous nucleic acids, and, more recently, have been implicated in bacterial virulence. Previously, our clinical findings identified activation of the CRISPR-Cas system in patient samples at the transition to disease; however, the mechanism of contribution to disease remained unknown. The importance of the present study resides in that it is becoming increasingly clear that CRISPR-associated proteins have broader functions than initially thought and that those functions now include their role in the virulence of periodontal pathogens. Studying a P. gingivalis cas3 mutant, we demonstrate that at least one of the CRISPR-Cas systems is involved in the regulation of virulence during infection.
format	article
author	Jose Solbiati Ana Duran-Pinedo Fernanda Godoy Rocha Frank C. Gibson Jorge Frias-Lopez
author_facet	Jose Solbiati Ana Duran-Pinedo Fernanda Godoy Rocha Frank C. Gibson Jorge Frias-Lopez
author_sort	Jose Solbiati
title	Virulence of the Pathogen <named-content content-type="genus-species">Porphyromonas gingivalis</named-content> Is Controlled by the CRISPR-Cas Protein Cas3
title_short	Virulence of the Pathogen <named-content content-type="genus-species">Porphyromonas gingivalis</named-content> Is Controlled by the CRISPR-Cas Protein Cas3
title_full	Virulence of the Pathogen <named-content content-type="genus-species">Porphyromonas gingivalis</named-content> Is Controlled by the CRISPR-Cas Protein Cas3
title_fullStr	Virulence of the Pathogen <named-content content-type="genus-species">Porphyromonas gingivalis</named-content> Is Controlled by the CRISPR-Cas Protein Cas3
title_full_unstemmed	Virulence of the Pathogen <named-content content-type="genus-species">Porphyromonas gingivalis</named-content> Is Controlled by the CRISPR-Cas Protein Cas3
title_sort	virulence of the pathogen <named-content content-type="genus-species">porphyromonas gingivalis</named-content> is controlled by the crispr-cas protein cas3
publisher	American Society for Microbiology
publishDate	2020
url	https://doaj.org/article/7bfecfb23ba64cbfa3d4bda9e35cc9bb
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Virulence of the Pathogen <named-content content-type="genus-species">Porphyromonas gingivalis</named-content> Is Controlled by the CRISPR-Cas Protein Cas3

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