The Passage of Chaperonins to Extracellular Locations in <i>Legionella pneumophila</i> Requires a Functional Dot/Icm System
HtpB, the chaperonin of the bacterial pathogen <i>L. pneumophila</i>, is found in extracellular locations, even the cytoplasm of host cells. Although chaperonins have an essential cytoplasmic function in protein folding, HtpB exits the cytoplasm to perform extracellular virulence-related...
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2025-01-01
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| author | Peter Robertson David S. Allan Rafael A. Garduño |
| author_facet | Peter Robertson David S. Allan Rafael A. Garduño |
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| description | HtpB, the chaperonin of the bacterial pathogen <i>L. pneumophila</i>, is found in extracellular locations, even the cytoplasm of host cells. Although chaperonins have an essential cytoplasmic function in protein folding, HtpB exits the cytoplasm to perform extracellular virulence-related functions that support <i>L. pneumophila</i>’s lifestyle. The mechanism by which HtpB reaches extracellular locations is not currently understood. To address this experimental gap, immunoelectron microscopy, trypsin-accessibility assays, and cell fractionation were used to localize HtpB in various <i>L. pneumophila</i> secretion mutants. Dot/Icm type IV secretion mutants displayed less surface-exposed HtpB and more periplasmic HtpB than parent strains. The analysis of periplasmic extracts and outer membrane vesicles of these mutants, where HtpB co-localized with bona fide periplasmic proteins, confirmed the elevated levels of periplasmic HtpB. Genetic complementation of the mutants recovered parent strain levels of surface-exposed and periplasmic HtpB. The export of GSK-tagged HtpB into the cytoplasm of infected cells was also Dot/Icm-dependent. The translocating role of the Dot/Icm system was not specific for HtpB because GroEL, the chaperonin of <i>Escherichia coli</i>, was found at the cell surface and accumulated in the periplasm of Dot mutants when expressed in <i>L. pneumophila</i>. These findings establish that a functional Dot/Icm system is required for HtpB to reach extracellular locations, but the mechanism by which cytoplasmic HtpB reaches the periplasm remains partially unidentified. |
| format | Article |
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| institution | Kabale University |
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| language | English |
| publishDate | 2025-01-01 |
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| spelling | doaj-art-d8145443b0434bc3925151527cd6eca92025-01-24T13:25:09ZengMDPI AGBiomolecules2218-273X2025-01-011519110.3390/biom15010091The Passage of Chaperonins to Extracellular Locations in <i>Legionella pneumophila</i> Requires a Functional Dot/Icm SystemPeter Robertson0David S. Allan1Rafael A. Garduño2Department of Microbiology-Immunology, Faculty of Medicine, Dalhousie University, Halifax, NS B3H 4R2, CanadaDepartment of Microbiology-Immunology, Faculty of Medicine, Dalhousie University, Halifax, NS B3H 4R2, CanadaDepartment of Microbiology-Immunology, Faculty of Medicine, Dalhousie University, Halifax, NS B3H 4R2, CanadaHtpB, the chaperonin of the bacterial pathogen <i>L. pneumophila</i>, is found in extracellular locations, even the cytoplasm of host cells. Although chaperonins have an essential cytoplasmic function in protein folding, HtpB exits the cytoplasm to perform extracellular virulence-related functions that support <i>L. pneumophila</i>’s lifestyle. The mechanism by which HtpB reaches extracellular locations is not currently understood. To address this experimental gap, immunoelectron microscopy, trypsin-accessibility assays, and cell fractionation were used to localize HtpB in various <i>L. pneumophila</i> secretion mutants. Dot/Icm type IV secretion mutants displayed less surface-exposed HtpB and more periplasmic HtpB than parent strains. The analysis of periplasmic extracts and outer membrane vesicles of these mutants, where HtpB co-localized with bona fide periplasmic proteins, confirmed the elevated levels of periplasmic HtpB. Genetic complementation of the mutants recovered parent strain levels of surface-exposed and periplasmic HtpB. The export of GSK-tagged HtpB into the cytoplasm of infected cells was also Dot/Icm-dependent. The translocating role of the Dot/Icm system was not specific for HtpB because GroEL, the chaperonin of <i>Escherichia coli</i>, was found at the cell surface and accumulated in the periplasm of Dot mutants when expressed in <i>L. pneumophila</i>. These findings establish that a functional Dot/Icm system is required for HtpB to reach extracellular locations, but the mechanism by which cytoplasmic HtpB reaches the periplasm remains partially unidentified.https://www.mdpi.com/2218-273X/15/1/91chaperoninsGroELHtpBHsp60 secretionDot/Icm systemimmunolabeling |
| spellingShingle | Peter Robertson David S. Allan Rafael A. Garduño The Passage of Chaperonins to Extracellular Locations in <i>Legionella pneumophila</i> Requires a Functional Dot/Icm System Biomolecules chaperonins GroEL HtpB Hsp60 secretion Dot/Icm system immunolabeling |
| title | The Passage of Chaperonins to Extracellular Locations in <i>Legionella pneumophila</i> Requires a Functional Dot/Icm System |
| title_full | The Passage of Chaperonins to Extracellular Locations in <i>Legionella pneumophila</i> Requires a Functional Dot/Icm System |
| title_fullStr | The Passage of Chaperonins to Extracellular Locations in <i>Legionella pneumophila</i> Requires a Functional Dot/Icm System |
| title_full_unstemmed | The Passage of Chaperonins to Extracellular Locations in <i>Legionella pneumophila</i> Requires a Functional Dot/Icm System |
| title_short | The Passage of Chaperonins to Extracellular Locations in <i>Legionella pneumophila</i> Requires a Functional Dot/Icm System |
| title_sort | passage of chaperonins to extracellular locations in i legionella pneumophila i requires a functional dot icm system |
| topic | chaperonins GroEL HtpB Hsp60 secretion Dot/Icm system immunolabeling |
| url | https://www.mdpi.com/2218-273X/15/1/91 |
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