Visualizing the Translocation and Localization of Bacterial Type III Effector Proteins by Using a Genetically Encoded Reporter System
Bacterial type III secretion system (T3SS) effector proteins are critical determinants of infection for many animal and plant pathogens. However, monitoring of the translocation and delivery of these important virulence determinants has proved to be technically challenging. Here, we used a genetical...
Gespeichert in:
| Veröffentlicht in: | Applied and environmental microbiology 2016-05, Vol.82 (9), p.2700-2708 |
|---|---|
| Hauptverfasser: | , , , , , , |
| Format: | Artikel |
| Sprache: | eng |
| Schlagworte: | |
| Online-Zugang: | Volltext |
| Tags: |
Tag hinzufügen
Keine Tags, Fügen Sie den ersten Tag hinzu!
|
| container_end_page | 2708 |
|---|---|
| container_issue | 9 |
| container_start_page | 2700 |
| container_title | Applied and environmental microbiology |
| container_volume | 82 |
| creator | Gawthorne, Jayde A Audry, Laurent McQuitty, Claire Dean, Paul Christie, John M Enninga, Jost Roe, Andrew J |
| description | Bacterial type III secretion system (T3SS) effector proteins are critical determinants of infection for many animal and plant pathogens. However, monitoring of the translocation and delivery of these important virulence determinants has proved to be technically challenging. Here, we used a genetically engineered LOV (light-oxygen-voltage) sensing domain derivative to monitor the expression, translocation, and localization of bacterial T3SS effectors. We found the Escherichia coli O157:H7 bacterial effector fusion Tir-LOV was functional following its translocation and localized to the host cell membrane in discrete foci, demonstrating that LOV-based reporters can be used to visualize the effector translocation with minimal manipulation and interference. Further evidence for the versatility of the reporter was demonstrated by fusing LOV to the C terminus of the Shigella flexneri effector IpaB. IpaB-LOV localized preferentially at bacterial poles before translocation. We observed the rapid translocation of IpaB-LOV in a T3SS-dependent manner into host cells, where it localized at the bacterial entry site within membrane ruffles. |
| doi_str_mv | 10.1128/AEM.03418-15 |
| format | Article |
| fullrecord | <record><control><sourceid>proquest_pubme</sourceid><recordid>TN_cdi_pubmedcentral_primary_oai_pubmedcentral_nih_gov_4836418</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><sourcerecordid>1782834731</sourcerecordid><originalsourceid>FETCH-LOGICAL-c498t-40eb105e2e911489b149d324f576923883ad7549cef576a88699d0d6b2ebf1f43</originalsourceid><addsrcrecordid>eNqNks9v0zAUxy0EYmVw44x85ECGfyWxL0hlKlulIhB0XC0neVmNUjvYzqRw3_89l44JOHGy_Pzx5_lZX4ReUnJGKZNvl6uPZ4QLKgtaPkILSpQsSs6rx2hBiFIFY4KcoGcxfieECFLJp-iEVYpRwaoFuv1m42QG-9O6a5x2gLfBuDj41iTrHTauw5u8ycCx4Hv83rQJgjUD3s4j4PV6jVd9D23yAX8OPoF1ETczvooHp8EX4CDZ7BhmvHKt76DDX2D0IVvw1zkm2D9HT3ozRHhxv56iqw-r7fllsfl0sT5fbopWKJkKQaChpAQGilIhVUOF6jgTfVnngbiU3HR1KVQLh4qRslKqI13VMGh62gt-it4dvePU7KFrwaVgBj0Guzdh1t5Y_feJszt97W-0kLzKX5wFxVGw--fa5XKjR5NnmYImVColBL-hmX993zD4HxPEpPc2tjAMxoGfoqa1IooLQuv_QCWTXNT8YH1zRNvgYwzQPzyFEn0Ihc6h0L9CoWmZ8Vd_Tv0A_04BvwMra7Ml</addsrcrecordid><sourcetype>Open Access Repository</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>1782834731</pqid></control><display><type>article</type><title>Visualizing the Translocation and Localization of Bacterial Type III Effector Proteins by Using a Genetically Encoded Reporter System</title><source>American Society for Microbiology</source><source>MEDLINE</source><source>PubMed Central</source><source>Alma/SFX Local Collection</source><creator>Gawthorne, Jayde A ; Audry, Laurent ; McQuitty, Claire ; Dean, Paul ; Christie, John M ; Enninga, Jost ; Roe, Andrew J</creator><contributor>Besser, T. E.</contributor><creatorcontrib>Gawthorne, Jayde A ; Audry, Laurent ; McQuitty, Claire ; Dean, Paul ; Christie, John M ; Enninga, Jost ; Roe, Andrew J ; Besser, T. E.</creatorcontrib><description>Bacterial type III secretion system (T3SS) effector proteins are critical determinants of infection for many animal and plant pathogens. However, monitoring of the translocation and delivery of these important virulence determinants has proved to be technically challenging. Here, we used a genetically engineered LOV (light-oxygen-voltage) sensing domain derivative to monitor the expression, translocation, and localization of bacterial T3SS effectors. We found the Escherichia coli O157:H7 bacterial effector fusion Tir-LOV was functional following its translocation and localized to the host cell membrane in discrete foci, demonstrating that LOV-based reporters can be used to visualize the effector translocation with minimal manipulation and interference. Further evidence for the versatility of the reporter was demonstrated by fusing LOV to the C terminus of the Shigella flexneri effector IpaB. IpaB-LOV localized preferentially at bacterial poles before translocation. We observed the rapid translocation of IpaB-LOV in a T3SS-dependent manner into host cells, where it localized at the bacterial entry site within membrane ruffles.</description><identifier>ISSN: 0099-2240</identifier><identifier>EISSN: 1098-5336</identifier><identifier>DOI: 10.1128/AEM.03418-15</identifier><identifier>PMID: 26921426</identifier><language>eng</language><publisher>United States: American Society for Microbiology</publisher><subject>Bacterial Proteins ; Bacterial Proteins - genetics ; Bacterial Proteins - metabolism ; Biochemistry ; Biochemistry, Molecular Biology ; Cell Behavior ; Cellular Biology ; Escherichia coli ; Escherichia coli O157 ; Escherichia coli O157 - genetics ; Escherichia coli O157 - metabolism ; Escherichia coli Proteins ; Escherichia coli Proteins - genetics ; Escherichia coli Proteins - metabolism ; Genes, Reporter ; Genetic Engineering ; Genetic Engineering - methods ; HeLa Cells ; Host-Pathogen Interactions ; Humans ; Life Sciences ; Methods ; Optical Imaging ; Protein Domains ; Shigella flexneri ; Shigella flexneri - genetics ; Shigella flexneri - metabolism ; Type III Secretion Systems ; Type III Secretion Systems - analysis ; Type III Secretion Systems - genetics ; Type III Secretion Systems - metabolism</subject><ispartof>Applied and environmental microbiology, 2016-05, Vol.82 (9), p.2700-2708</ispartof><rights>Copyright © 2016 Gawthorne et al.</rights><rights>Attribution</rights><rights>Copyright © 2016 Gawthorne et al. 2016 Gawthorne et al.</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c498t-40eb105e2e911489b149d324f576923883ad7549cef576a88699d0d6b2ebf1f43</citedby><cites>FETCH-LOGICAL-c498t-40eb105e2e911489b149d324f576923883ad7549cef576a88699d0d6b2ebf1f43</cites><orcidid>0000-0003-3698-6134</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC4836418/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC4836418/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,723,776,780,881,3174,27903,27904,53769,53771</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/26921426$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink><backlink>$$Uhttps://pasteur.hal.science/pasteur-01899443$$DView record in HAL$$Hfree_for_read</backlink></links><search><contributor>Besser, T. E.</contributor><creatorcontrib>Gawthorne, Jayde A</creatorcontrib><creatorcontrib>Audry, Laurent</creatorcontrib><creatorcontrib>McQuitty, Claire</creatorcontrib><creatorcontrib>Dean, Paul</creatorcontrib><creatorcontrib>Christie, John M</creatorcontrib><creatorcontrib>Enninga, Jost</creatorcontrib><creatorcontrib>Roe, Andrew J</creatorcontrib><title>Visualizing the Translocation and Localization of Bacterial Type III Effector Proteins by Using a Genetically Encoded Reporter System</title><title>Applied and environmental microbiology</title><addtitle>Appl Environ Microbiol</addtitle><description>Bacterial type III secretion system (T3SS) effector proteins are critical determinants of infection for many animal and plant pathogens. However, monitoring of the translocation and delivery of these important virulence determinants has proved to be technically challenging. Here, we used a genetically engineered LOV (light-oxygen-voltage) sensing domain derivative to monitor the expression, translocation, and localization of bacterial T3SS effectors. We found the Escherichia coli O157:H7 bacterial effector fusion Tir-LOV was functional following its translocation and localized to the host cell membrane in discrete foci, demonstrating that LOV-based reporters can be used to visualize the effector translocation with minimal manipulation and interference. Further evidence for the versatility of the reporter was demonstrated by fusing LOV to the C terminus of the Shigella flexneri effector IpaB. IpaB-LOV localized preferentially at bacterial poles before translocation. We observed the rapid translocation of IpaB-LOV in a T3SS-dependent manner into host cells, where it localized at the bacterial entry site within membrane ruffles.</description><subject>Bacterial Proteins</subject><subject>Bacterial Proteins - genetics</subject><subject>Bacterial Proteins - metabolism</subject><subject>Biochemistry</subject><subject>Biochemistry, Molecular Biology</subject><subject>Cell Behavior</subject><subject>Cellular Biology</subject><subject>Escherichia coli</subject><subject>Escherichia coli O157</subject><subject>Escherichia coli O157 - genetics</subject><subject>Escherichia coli O157 - metabolism</subject><subject>Escherichia coli Proteins</subject><subject>Escherichia coli Proteins - genetics</subject><subject>Escherichia coli Proteins - metabolism</subject><subject>Genes, Reporter</subject><subject>Genetic Engineering</subject><subject>Genetic Engineering - methods</subject><subject>HeLa Cells</subject><subject>Host-Pathogen Interactions</subject><subject>Humans</subject><subject>Life Sciences</subject><subject>Methods</subject><subject>Optical Imaging</subject><subject>Protein Domains</subject><subject>Shigella flexneri</subject><subject>Shigella flexneri - genetics</subject><subject>Shigella flexneri - metabolism</subject><subject>Type III Secretion Systems</subject><subject>Type III Secretion Systems - analysis</subject><subject>Type III Secretion Systems - genetics</subject><subject>Type III Secretion Systems - metabolism</subject><issn>0099-2240</issn><issn>1098-5336</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2016</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqNks9v0zAUxy0EYmVw44x85ECGfyWxL0hlKlulIhB0XC0neVmNUjvYzqRw3_89l44JOHGy_Pzx5_lZX4ReUnJGKZNvl6uPZ4QLKgtaPkILSpQsSs6rx2hBiFIFY4KcoGcxfieECFLJp-iEVYpRwaoFuv1m42QG-9O6a5x2gLfBuDj41iTrHTauw5u8ycCx4Hv83rQJgjUD3s4j4PV6jVd9D23yAX8OPoF1ETczvooHp8EX4CDZ7BhmvHKt76DDX2D0IVvw1zkm2D9HT3ozRHhxv56iqw-r7fllsfl0sT5fbopWKJkKQaChpAQGilIhVUOF6jgTfVnngbiU3HR1KVQLh4qRslKqI13VMGh62gt-it4dvePU7KFrwaVgBj0Guzdh1t5Y_feJszt97W-0kLzKX5wFxVGw--fa5XKjR5NnmYImVColBL-hmX993zD4HxPEpPc2tjAMxoGfoqa1IooLQuv_QCWTXNT8YH1zRNvgYwzQPzyFEn0Ihc6h0L9CoWmZ8Vd_Tv0A_04BvwMra7Ml</recordid><startdate>20160501</startdate><enddate>20160501</enddate><creator>Gawthorne, Jayde A</creator><creator>Audry, Laurent</creator><creator>McQuitty, Claire</creator><creator>Dean, Paul</creator><creator>Christie, John M</creator><creator>Enninga, Jost</creator><creator>Roe, Andrew J</creator><general>American Society for Microbiology</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope><scope>7QL</scope><scope>7T7</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>P64</scope><scope>1XC</scope><scope>VOOES</scope><scope>5PM</scope><orcidid>https://orcid.org/0000-0003-3698-6134</orcidid></search><sort><creationdate>20160501</creationdate><title>Visualizing the Translocation and Localization of Bacterial Type III Effector Proteins by Using a Genetically Encoded Reporter System</title><author>Gawthorne, Jayde A ; Audry, Laurent ; McQuitty, Claire ; Dean, Paul ; Christie, John M ; Enninga, Jost ; Roe, Andrew J</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c498t-40eb105e2e911489b149d324f576923883ad7549cef576a88699d0d6b2ebf1f43</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2016</creationdate><topic>Bacterial Proteins</topic><topic>Bacterial Proteins - genetics</topic><topic>Bacterial Proteins - metabolism</topic><topic>Biochemistry</topic><topic>Biochemistry, Molecular Biology</topic><topic>Cell Behavior</topic><topic>Cellular Biology</topic><topic>Escherichia coli</topic><topic>Escherichia coli O157</topic><topic>Escherichia coli O157 - genetics</topic><topic>Escherichia coli O157 - metabolism</topic><topic>Escherichia coli Proteins</topic><topic>Escherichia coli Proteins - genetics</topic><topic>Escherichia coli Proteins - metabolism</topic><topic>Genes, Reporter</topic><topic>Genetic Engineering</topic><topic>Genetic Engineering - methods</topic><topic>HeLa Cells</topic><topic>Host-Pathogen Interactions</topic><topic>Humans</topic><topic>Life Sciences</topic><topic>Methods</topic><topic>Optical Imaging</topic><topic>Protein Domains</topic><topic>Shigella flexneri</topic><topic>Shigella flexneri - genetics</topic><topic>Shigella flexneri - metabolism</topic><topic>Type III Secretion Systems</topic><topic>Type III Secretion Systems - analysis</topic><topic>Type III Secretion Systems - genetics</topic><topic>Type III Secretion Systems - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Gawthorne, Jayde A</creatorcontrib><creatorcontrib>Audry, Laurent</creatorcontrib><creatorcontrib>McQuitty, Claire</creatorcontrib><creatorcontrib>Dean, Paul</creatorcontrib><creatorcontrib>Christie, John M</creatorcontrib><creatorcontrib>Enninga, Jost</creatorcontrib><creatorcontrib>Roe, Andrew J</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Industrial and Applied Microbiology Abstracts (Microbiology A)</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>Engineering Research Database</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Hyper Article en Ligne (HAL)</collection><collection>Hyper Article en Ligne (HAL) (Open Access)</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Applied and environmental microbiology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Gawthorne, Jayde A</au><au>Audry, Laurent</au><au>McQuitty, Claire</au><au>Dean, Paul</au><au>Christie, John M</au><au>Enninga, Jost</au><au>Roe, Andrew J</au><au>Besser, T. E.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Visualizing the Translocation and Localization of Bacterial Type III Effector Proteins by Using a Genetically Encoded Reporter System</atitle><jtitle>Applied and environmental microbiology</jtitle><addtitle>Appl Environ Microbiol</addtitle><date>2016-05-01</date><risdate>2016</risdate><volume>82</volume><issue>9</issue><spage>2700</spage><epage>2708</epage><pages>2700-2708</pages><issn>0099-2240</issn><eissn>1098-5336</eissn><abstract>Bacterial type III secretion system (T3SS) effector proteins are critical determinants of infection for many animal and plant pathogens. However, monitoring of the translocation and delivery of these important virulence determinants has proved to be technically challenging. Here, we used a genetically engineered LOV (light-oxygen-voltage) sensing domain derivative to monitor the expression, translocation, and localization of bacterial T3SS effectors. We found the Escherichia coli O157:H7 bacterial effector fusion Tir-LOV was functional following its translocation and localized to the host cell membrane in discrete foci, demonstrating that LOV-based reporters can be used to visualize the effector translocation with minimal manipulation and interference. Further evidence for the versatility of the reporter was demonstrated by fusing LOV to the C terminus of the Shigella flexneri effector IpaB. IpaB-LOV localized preferentially at bacterial poles before translocation. We observed the rapid translocation of IpaB-LOV in a T3SS-dependent manner into host cells, where it localized at the bacterial entry site within membrane ruffles.</abstract><cop>United States</cop><pub>American Society for Microbiology</pub><pmid>26921426</pmid><doi>10.1128/AEM.03418-15</doi><tpages>9</tpages><orcidid>https://orcid.org/0000-0003-3698-6134</orcidid><oa>free_for_read</oa></addata></record> |
| fulltext | fulltext |
| identifier | ISSN: 0099-2240 |
| ispartof | Applied and environmental microbiology, 2016-05, Vol.82 (9), p.2700-2708 |
| issn | 0099-2240 1098-5336 |
| language | eng |
| recordid | cdi_pubmedcentral_primary_oai_pubmedcentral_nih_gov_4836418 |
| source | American Society for Microbiology; MEDLINE; PubMed Central; Alma/SFX Local Collection |
| subjects | Bacterial Proteins Bacterial Proteins - genetics Bacterial Proteins - metabolism Biochemistry Biochemistry, Molecular Biology Cell Behavior Cellular Biology Escherichia coli Escherichia coli O157 Escherichia coli O157 - genetics Escherichia coli O157 - metabolism Escherichia coli Proteins Escherichia coli Proteins - genetics Escherichia coli Proteins - metabolism Genes, Reporter Genetic Engineering Genetic Engineering - methods HeLa Cells Host-Pathogen Interactions Humans Life Sciences Methods Optical Imaging Protein Domains Shigella flexneri Shigella flexneri - genetics Shigella flexneri - metabolism Type III Secretion Systems Type III Secretion Systems - analysis Type III Secretion Systems - genetics Type III Secretion Systems - metabolism |
| title | Visualizing the Translocation and Localization of Bacterial Type III Effector Proteins by Using a Genetically Encoded Reporter System |
| url | https://sfx.bib-bvb.de/sfx_tum?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2025-01-23T22%3A51%3A08IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-proquest_pubme&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=Visualizing%20the%20Translocation%20and%20Localization%20of%20Bacterial%20Type%20III%20Effector%20Proteins%20by%20Using%20a%20Genetically%20Encoded%20Reporter%20System&rft.jtitle=Applied%20and%20environmental%20microbiology&rft.au=Gawthorne,%20Jayde%20A&rft.date=2016-05-01&rft.volume=82&rft.issue=9&rft.spage=2700&rft.epage=2708&rft.pages=2700-2708&rft.issn=0099-2240&rft.eissn=1098-5336&rft_id=info:doi/10.1128/AEM.03418-15&rft_dat=%3Cproquest_pubme%3E1782834731%3C/proquest_pubme%3E%3Curl%3E%3C/url%3E&disable_directlink=true&sfx.directlink=off&sfx.report_link=0&rft_id=info:oai/&rft_pqid=1782834731&rft_id=info:pmid/26921426&rfr_iscdi=true |