Flagellar Hook Protein FlgE Induces Microvascular Hyperpermeability via Ectopic ATP Synthase β on Endothelial Surface
We previously demonstrated the immunostimulatory efficacy of Pseudomonas aeruginosa flagellar hook protein FlgE on epithelial cells, presumably via ectopic ATP synthases or subunits ATP5B on cell membranes. Here, by using recombinant wild-type FlgE, mutant FlgE (FlgEM; bearing mutations on two postu...
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Frontiers Media S.A.
2021-11-01
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| Series: | Frontiers in Cellular and Infection Microbiology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fcimb.2021.724912/full |
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| author | Yuanyuan Li Yuanyuan Li Ying Shen Yudan Zheng Shundong Ji Mengru Wang Beibei Wang Qingzhen Han Yufeng Tian Yiqiang Wang Yiqiang Wang |
| author_facet | Yuanyuan Li Yuanyuan Li Ying Shen Yudan Zheng Shundong Ji Mengru Wang Beibei Wang Qingzhen Han Yufeng Tian Yiqiang Wang Yiqiang Wang |
| author_sort | Yuanyuan Li |
| collection | DOAJ |
| description | We previously demonstrated the immunostimulatory efficacy of Pseudomonas aeruginosa flagellar hook protein FlgE on epithelial cells, presumably via ectopic ATP synthases or subunits ATP5B on cell membranes. Here, by using recombinant wild-type FlgE, mutant FlgE (FlgEM; bearing mutations on two postulated critical epitopes B and F), and a FlgE analog in pull-down assay, Western blotting, flow cytometry, and ELISA, actual bindings of FlgE proteins or epitope B/F peptides with ATP5B were all confirmed. Upon treatment with FlgE proteins, human umbilical vein endothelial cells (HUVECs) and SV40-immortalized murine vascular endothelial cells manifested decreased proliferation, migration, tube formation, and surface ATP production and increased apoptosis. FlgE proteins increased the permeability of HUVEC monolayers to soluble large molecules like dextran as well as to neutrophils. Immunofluorescence showed that FlgE induced clustering and conjugation of F-actin in HUVECs. In Balb/c-nude mice bearing transplanted solid tumors, FlgE proteins induced a microvascular hyperpermeability in pinna, lungs, tumor mass, and abdominal cavity. All effects observed in FlgE proteins were partially or completely impaired in FlgEM proteins or blocked by pretreatment with anti-ATP5B antibodies. Upon coculture of bacteria with HUVECs, FlgE was detectable in the membrane and cytosol of HUVECs. It was concluded that FlgE posed a pathogenic ligand of ectopic ATP5B that, upon FlgE–ATP5B coupling on endothelial cells, modulated properties and increased permeability of endothelial layers both in vitro and in vivo. The FlgE-ectopic ATP5B duo might contribute to the pathogenesis of disorders associated with bacterial infection or ectopic ATP5B-positive cells. |
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| language | English |
| last_indexed | 2024-12-17T13:53:27Z |
| publishDate | 2021-11-01 |
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| spelling | doaj.art-cb4cc1c84c13432e9ca6af884769b8dc2022-12-21T21:45:59ZengFrontiers Media S.A.Frontiers in Cellular and Infection Microbiology2235-29882021-11-011110.3389/fcimb.2021.724912724912Flagellar Hook Protein FlgE Induces Microvascular Hyperpermeability via Ectopic ATP Synthase β on Endothelial SurfaceYuanyuan Li0Yuanyuan Li1Ying Shen2Yudan Zheng3Shundong Ji4Mengru Wang5Beibei Wang6Qingzhen Han7Yufeng Tian8Yiqiang Wang9Yiqiang Wang10The MOH Key Lab of Thrombosis and Hemostasis, The First Affiliated Hospital of Soochow University, Soochow University, Suzhou, ChinaDepartment of Laboratory Examination, People’s Hospital of Rizhao City, Rizhao, ChinaThe MOH Key Lab of Thrombosis and Hemostasis, The First Affiliated Hospital of Soochow University, Soochow University, Suzhou, ChinaThe MOH Key Lab of Thrombosis and Hemostasis, The First Affiliated Hospital of Soochow University, Soochow University, Suzhou, ChinaThe MOH Key Lab of Thrombosis and Hemostasis, The First Affiliated Hospital of Soochow University, Soochow University, Suzhou, ChinaThe MOH Key Lab of Thrombosis and Hemostasis, The First Affiliated Hospital of Soochow University, Soochow University, Suzhou, ChinaCenter for Informational Biology, School of Life Science and Technology, University of Electronic Science and Technology of China, Chengdu, ChinaDepartment of Laboratory Examination, The First Affiliated Hospital of Soochow University, Soochow University, Suzhou, ChinaDepartment of Laboratory Examination, People’s Hospital of Rizhao City, Rizhao, ChinaThe MOH Key Lab of Thrombosis and Hemostasis, The First Affiliated Hospital of Soochow University, Soochow University, Suzhou, ChinaCentral Lab, Xiang’an Hospital of Xiamen University, Xiamen University Medical Center, Xiamen University, Xiamen, ChinaWe previously demonstrated the immunostimulatory efficacy of Pseudomonas aeruginosa flagellar hook protein FlgE on epithelial cells, presumably via ectopic ATP synthases or subunits ATP5B on cell membranes. Here, by using recombinant wild-type FlgE, mutant FlgE (FlgEM; bearing mutations on two postulated critical epitopes B and F), and a FlgE analog in pull-down assay, Western blotting, flow cytometry, and ELISA, actual bindings of FlgE proteins or epitope B/F peptides with ATP5B were all confirmed. Upon treatment with FlgE proteins, human umbilical vein endothelial cells (HUVECs) and SV40-immortalized murine vascular endothelial cells manifested decreased proliferation, migration, tube formation, and surface ATP production and increased apoptosis. FlgE proteins increased the permeability of HUVEC monolayers to soluble large molecules like dextran as well as to neutrophils. Immunofluorescence showed that FlgE induced clustering and conjugation of F-actin in HUVECs. In Balb/c-nude mice bearing transplanted solid tumors, FlgE proteins induced a microvascular hyperpermeability in pinna, lungs, tumor mass, and abdominal cavity. All effects observed in FlgE proteins were partially or completely impaired in FlgEM proteins or blocked by pretreatment with anti-ATP5B antibodies. Upon coculture of bacteria with HUVECs, FlgE was detectable in the membrane and cytosol of HUVECs. It was concluded that FlgE posed a pathogenic ligand of ectopic ATP5B that, upon FlgE–ATP5B coupling on endothelial cells, modulated properties and increased permeability of endothelial layers both in vitro and in vivo. The FlgE-ectopic ATP5B duo might contribute to the pathogenesis of disorders associated with bacterial infection or ectopic ATP5B-positive cells.https://www.frontiersin.org/articles/10.3389/fcimb.2021.724912/fullATP synthase subunit βATP5Bflagellar hook proteinFlgEvascular endothelial cells |
| spellingShingle | Yuanyuan Li Yuanyuan Li Ying Shen Yudan Zheng Shundong Ji Mengru Wang Beibei Wang Qingzhen Han Yufeng Tian Yiqiang Wang Yiqiang Wang Flagellar Hook Protein FlgE Induces Microvascular Hyperpermeability via Ectopic ATP Synthase β on Endothelial Surface Frontiers in Cellular and Infection Microbiology ATP synthase subunit β ATP5B flagellar hook protein FlgE vascular endothelial cells |
| title | Flagellar Hook Protein FlgE Induces Microvascular Hyperpermeability via Ectopic ATP Synthase β on Endothelial Surface |
| title_full | Flagellar Hook Protein FlgE Induces Microvascular Hyperpermeability via Ectopic ATP Synthase β on Endothelial Surface |
| title_fullStr | Flagellar Hook Protein FlgE Induces Microvascular Hyperpermeability via Ectopic ATP Synthase β on Endothelial Surface |
| title_full_unstemmed | Flagellar Hook Protein FlgE Induces Microvascular Hyperpermeability via Ectopic ATP Synthase β on Endothelial Surface |
| title_short | Flagellar Hook Protein FlgE Induces Microvascular Hyperpermeability via Ectopic ATP Synthase β on Endothelial Surface |
| title_sort | flagellar hook protein flge induces microvascular hyperpermeability via ectopic atp synthase β on endothelial surface |
| topic | ATP synthase subunit β ATP5B flagellar hook protein FlgE vascular endothelial cells |
| url | https://www.frontiersin.org/articles/10.3389/fcimb.2021.724912/full |
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