Unraveling and characterization of novel T3SS effectors in Edwardsiella piscicida
ABSTRACT Type III secretion system (T3SS) facilitates survival and replication of Edwardsiella piscicida in vivo. Identifying novel T3SS effectors and elucidating their functions are critical in understanding the pathogenesis of E. piscicida. E. piscicida T3SS effector EseG and EseJ was highly secre...
| Main Authors: | , , , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
American Society for Microbiology
2023-10-01
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| Series: | mSphere |
| Subjects: | |
| Online Access: | https://journals.asm.org/doi/10.1128/msphere.00346-23 |
| _version_ | 1797651074165768192 |
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| author | Xiao Jian Liao Tian Tian He Lu Yi Liu Xiu Long Jiang Shan Shan Sun Yu Hang Deng Li Qiang Zhang Hai Xia Xie Pin Nie |
| author_facet | Xiao Jian Liao Tian Tian He Lu Yi Liu Xiu Long Jiang Shan Shan Sun Yu Hang Deng Li Qiang Zhang Hai Xia Xie Pin Nie |
| author_sort | Xiao Jian Liao |
| collection | DOAJ |
| description | ABSTRACT Type III secretion system (T3SS) facilitates survival and replication of Edwardsiella piscicida in vivo. Identifying novel T3SS effectors and elucidating their functions are critical in understanding the pathogenesis of E. piscicida. E. piscicida T3SS effector EseG and EseJ was highly secreted when T3SS gatekeeper-containing protein complex EsaB-EsaL-EsaM was disrupted by EsaB deficiency. Based on this observation, concentrated secretomes of ΔesaB strain and ΔesaBΔesaN strain were purified by loading them into SDS-PAGE gel for a short electrophoresis to remove impurities prior to the in-the gel digestion and mass spectrometry. Four reported T3SS effectors and two novel T3SS effector candidates EseQ (ETAE_2009) and Trx2 (ETAE_0559) were unraveled by quantitative comparison of the identified peptides. EseQ and Trx2 were revealed to be secreted and translocated in a T3SS-dependent manner through CyaA-based translocation assay and immunofluorescent staining, demonstrating that EseQ and Trx2 are the novel T3SS effectors of E. piscicida. Trx2 was found to suppress macrophage apoptosis as revealed by TUNEL staining and cleaved caspase-3 of infected J774A.1 monolayers. Moreover, Trx2 has been shown to inhibit the p65 phosphorylation and p65 translocation into the nucleus, thus blocking the NF-κB pathway. Furthermore, depletion of Trx2 slightly but significantly attenuates E. piscicida virulence in a fish infection model. Taken together, an efficient method was established in unraveling T3SS effectors in E. piscicida, and Trx2, one of the novel T3SS effectors identified in this study, was demonstrated to suppress apoptosis and block NF- κB pathway during E. piscicida infection. IMPORTANCE Edwardsiella piscicida is an intracellular bacterial pathogen that causes intestinal inflammation and hemorrhagic sepsis in fish and human. Virulence depends on the Edwardsiella type III secretion system (T3SS). Identifying the bacterial effector proteins secreted by T3SS and defining their role is key to understanding Edwardsiella pathogenesis. EsaB depletion disrupts the T3SS gatekeeper-containing protein complex, resulting in increased secretion of T3SS effectors EseG and EseJ. EseQ and Trx2 were shown to be the novel T3SS effectors of E. piscicida by a secretome comparison between ∆esaB strain and ∆esaB∆esaN strain (T3SS mutant), together with CyaA-based translocation assay. In addition, Trx2 has been shown to suppress macrophage apoptosis and block the NF-κB pathway. Together, this work expands the known repertoire of T3SS effectors and sheds light on the pathogenic mechanism of E. piscicida. |
| first_indexed | 2024-03-11T16:09:47Z |
| format | Article |
| id | doaj.art-f5a5a310835a4fc5b2178cec14f31174 |
| institution | Directory Open Access Journal |
| issn | 2379-5042 |
| language | English |
| last_indexed | 2024-03-11T16:09:47Z |
| publishDate | 2023-10-01 |
| publisher | American Society for Microbiology |
| record_format | Article |
| series | mSphere |
| spelling | doaj.art-f5a5a310835a4fc5b2178cec14f311742023-10-24T16:32:15ZengAmerican Society for MicrobiologymSphere2379-50422023-10-018510.1128/msphere.00346-23Unraveling and characterization of novel T3SS effectors in Edwardsiella piscicidaXiao Jian Liao0Tian Tian He1Lu Yi Liu2Xiu Long Jiang3Shan Shan Sun4Yu Hang Deng5Li Qiang Zhang6Hai Xia Xie7Pin Nie8State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences , Wuhan, ChinaState Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences , Wuhan, ChinaState Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences , Wuhan, ChinaState Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences , Wuhan, ChinaState Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences , Wuhan, ChinaState Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences , Wuhan, ChinaFisheries Research Institute, Wuhan Academy of Agricultural Sciences , Wuhan, ChinaState Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences , Wuhan, ChinaState Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences , Wuhan, ChinaABSTRACT Type III secretion system (T3SS) facilitates survival and replication of Edwardsiella piscicida in vivo. Identifying novel T3SS effectors and elucidating their functions are critical in understanding the pathogenesis of E. piscicida. E. piscicida T3SS effector EseG and EseJ was highly secreted when T3SS gatekeeper-containing protein complex EsaB-EsaL-EsaM was disrupted by EsaB deficiency. Based on this observation, concentrated secretomes of ΔesaB strain and ΔesaBΔesaN strain were purified by loading them into SDS-PAGE gel for a short electrophoresis to remove impurities prior to the in-the gel digestion and mass spectrometry. Four reported T3SS effectors and two novel T3SS effector candidates EseQ (ETAE_2009) and Trx2 (ETAE_0559) were unraveled by quantitative comparison of the identified peptides. EseQ and Trx2 were revealed to be secreted and translocated in a T3SS-dependent manner through CyaA-based translocation assay and immunofluorescent staining, demonstrating that EseQ and Trx2 are the novel T3SS effectors of E. piscicida. Trx2 was found to suppress macrophage apoptosis as revealed by TUNEL staining and cleaved caspase-3 of infected J774A.1 monolayers. Moreover, Trx2 has been shown to inhibit the p65 phosphorylation and p65 translocation into the nucleus, thus blocking the NF-κB pathway. Furthermore, depletion of Trx2 slightly but significantly attenuates E. piscicida virulence in a fish infection model. Taken together, an efficient method was established in unraveling T3SS effectors in E. piscicida, and Trx2, one of the novel T3SS effectors identified in this study, was demonstrated to suppress apoptosis and block NF- κB pathway during E. piscicida infection. IMPORTANCE Edwardsiella piscicida is an intracellular bacterial pathogen that causes intestinal inflammation and hemorrhagic sepsis in fish and human. Virulence depends on the Edwardsiella type III secretion system (T3SS). Identifying the bacterial effector proteins secreted by T3SS and defining their role is key to understanding Edwardsiella pathogenesis. EsaB depletion disrupts the T3SS gatekeeper-containing protein complex, resulting in increased secretion of T3SS effectors EseG and EseJ. EseQ and Trx2 were shown to be the novel T3SS effectors of E. piscicida by a secretome comparison between ∆esaB strain and ∆esaB∆esaN strain (T3SS mutant), together with CyaA-based translocation assay. In addition, Trx2 has been shown to suppress macrophage apoptosis and block the NF-κB pathway. Together, this work expands the known repertoire of T3SS effectors and sheds light on the pathogenic mechanism of E. piscicida.https://journals.asm.org/doi/10.1128/msphere.00346-23comparative proteomicssecretomeT3SS effectorsEdwardsiella piscicida |
| spellingShingle | Xiao Jian Liao Tian Tian He Lu Yi Liu Xiu Long Jiang Shan Shan Sun Yu Hang Deng Li Qiang Zhang Hai Xia Xie Pin Nie Unraveling and characterization of novel T3SS effectors in Edwardsiella piscicida mSphere comparative proteomics secretome T3SS effectors Edwardsiella piscicida |
| title | Unraveling and characterization of novel T3SS effectors in Edwardsiella piscicida |
| title_full | Unraveling and characterization of novel T3SS effectors in Edwardsiella piscicida |
| title_fullStr | Unraveling and characterization of novel T3SS effectors in Edwardsiella piscicida |
| title_full_unstemmed | Unraveling and characterization of novel T3SS effectors in Edwardsiella piscicida |
| title_short | Unraveling and characterization of novel T3SS effectors in Edwardsiella piscicida |
| title_sort | unraveling and characterization of novel t3ss effectors in edwardsiella piscicida |
| topic | comparative proteomics secretome T3SS effectors Edwardsiella piscicida |
| url | https://journals.asm.org/doi/10.1128/msphere.00346-23 |
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