[{"user_id":"317138e5-6ab7-11ef-aa6d-ffef3953e345","date_created":"2024-04-19T09:54:59Z","doi":"10.1242/jcs.261720","acknowledgement":"Nataliia Gnyliukh was partially funded by the European Union’s Horizon 2020 research and\r\ninnovation program (2018-2020) under the Marie Sklodowska-Curie Grant (agreement no.\r\n665385). Taif University Researchers Supporting Project: TURSP-HC2022/02. and Austrian\r\nScience Fund (FWF): I 6123-B.We thank Prof. Eileen Lafer and Liping Wang for their suggestions regarding the optimisation of protein expression and purification. We thank Prof. Sebastian Y. Bednarek for the useful comments and constructive criticism of the project. We thank Maciek Adamowski for providing genetic material. This research was supported by the Scientific Service Units (SSU) of IST-Austria through resources provided by the Electron microscopy (EMF), Lab Support Facility (LSF) (particularly Dorota Jaworska) and the Bioimaging Facility (BIF).","article_number":"jcs.261720","year":"2024","publication_identifier":{"issn":["0021-9533"],"eissn":["1477-9137"]},"publication_status":"published","article_type":"original","project":[{"call_identifier":"H2020","_id":"2564DBCA-B435-11E9-9278-68D0E5697425","grant_number":"665385","name":"International IST Doctoral Program"},{"grant_number":"I06123","name":"Peptide receptors for auxin canalization in Arabidopsis","_id":"bd76d395-d553-11ed-ba76-f678c14f9033"}],"type":"journal_article","title":"Role of dynamin-related proteins 2 and SH3P2 in clathrin-mediated endocytosis in Arabidopsis thaliana","language":[{"iso":"eng"}],"isi":1,"pmid":1,"oa_version":"Published Version","issue":"8","author":[{"orcid":"0000-0002-2198-0509","first_name":"Nataliia","id":"390C1120-F248-11E8-B48F-1D18A9856A87","full_name":"Gnyliukh, Nataliia","last_name":"Gnyliukh"},{"first_name":"Alexander J","id":"46A62C3A-F248-11E8-B48F-1D18A9856A87","full_name":"Johnson, Alexander J","last_name":"Johnson","orcid":"0000-0002-2739-8843"},{"first_name":"MK","full_name":"Nagel, MK","last_name":"Nagel"},{"id":"2DB5D88C-D7B3-11E9-B8FD-7907E6697425","first_name":"Aline","last_name":"Monzer","full_name":"Monzer, Aline"},{"last_name":"Babic","full_name":"Babic, David","id":"db566d23-f6e0-11ea-865d-e6f270e968e7","first_name":"David"},{"full_name":"Hlavata, Annamaria","last_name":"Hlavata","first_name":"Annamaria","id":"36062FEC-F248-11E8-B48F-1D18A9856A87"},{"full_name":"Alotaibi, SS","last_name":"Alotaibi","first_name":"SS"},{"full_name":"Isono, E","last_name":"Isono","first_name":"E"},{"orcid":"0000-0001-7309-9724","last_name":"Loose","full_name":"Loose, Martin","id":"462D4284-F248-11E8-B48F-1D18A9856A87","first_name":"Martin"},{"orcid":"0000-0002-8302-7596","full_name":"Friml, Jiří","last_name":"Friml","first_name":"Jiří","id":"4159519E-F248-11E8-B48F-1D18A9856A87"}],"day":"01","publication":"Journal of Cell Science","date_published":"2024-04-01T00:00:00Z","external_id":{"pmid":["38506228"],"isi":["001266917100005"]},"file":[{"checksum":"6dc023f0cc7052ad3cf0a42589d2e30f","relation":"main_file","access_level":"open_access","file_id":"18792","file_name":"2024_JourCellScience_Gnyliukh.pdf","date_updated":"2025-01-09T08:41:16Z","creator":"dernst","file_size":25845948,"date_created":"2025-01-09T08:41:16Z","content_type":"application/pdf","success":1}],"abstract":[{"lang":"eng","text":"Clathrin-mediated endocytosis (CME) is vital for the regulation of plant growth and development by controlling plasma membrane protein composition and cargo uptake. CME relies on the precise recruitment of regulators for vesicle maturation and release. Homologues of components of mammalian vesicle scission are strong candidates to be part of the scission machinery in plants, but the precise roles of these proteins in this process are not fully understood. Here, we characterised the roles of Plant Dynamin-Related Proteins 2 (DRP2s) and SH3-domain containing protein 2 (SH3P2), the plant homologue to Dynamins’ recruiters, like Endophilin and Amphiphysin, in the CME by combining high-resolution imaging of endocytic events in vivo and characterisation of the purified proteins in vitro. Although DRP2s and SH3P2 arrive similarly late during CME and physically interact, genetic analysis of the sh3p123 triple-mutant and complementation assays with non-SH3P2-interacting DRP2 variants suggests that SH3P2 does not directly recruit DRP2s to the site of endocytosis. These observations imply that despite the presence of many well-conserved endocytic components, plants have acquired a distinct mechanism for CME."}],"oa":1,"related_material":{"record":[{"relation":"earlier_version","status":"public","id":"14591"}]},"date_updated":"2025-09-04T13:49:45Z","status":"public","_id":"15330","corr_author":"1","scopus_import":"1","ddc":["570"],"file_date_updated":"2025-01-09T08:41:16Z","department":[{"_id":"MaLo"},{"_id":"JiFr"},{"_id":"CaBe"}],"quality_controlled":"1","volume":137,"OA_type":"hybrid","has_accepted_license":"1","citation":{"short":"N. Gnyliukh, A.J. Johnson, M. Nagel, A. Monzer, D. Babic, A. Hlavata, S. Alotaibi, E. Isono, M. Loose, J. Friml, Journal of Cell Science 137 (2024).","ista":"Gnyliukh N, Johnson AJ, Nagel M, Monzer A, Babic D, Hlavata A, Alotaibi S, Isono E, Loose M, Friml J. 2024. Role of dynamin-related proteins 2 and SH3P2 in clathrin-mediated endocytosis in Arabidopsis thaliana. Journal of Cell Science. 137(8), jcs. 261720.","mla":"Gnyliukh, Nataliia, et al. “Role of Dynamin-Related Proteins 2 and SH3P2 in Clathrin-Mediated Endocytosis in Arabidopsis Thaliana.” <i>Journal of Cell Science</i>, vol. 137, no. 8, jcs. 261720, The Company of Biologists, 2024, doi:<a href=\"https://doi.org/10.1242/jcs.261720\">10.1242/jcs.261720</a>.","apa":"Gnyliukh, N., Johnson, A. J., Nagel, M., Monzer, A., Babic, D., Hlavata, A., … Friml, J. (2024). Role of dynamin-related proteins 2 and SH3P2 in clathrin-mediated endocytosis in Arabidopsis thaliana. <i>Journal of Cell Science</i>. The Company of Biologists. <a href=\"https://doi.org/10.1242/jcs.261720\">https://doi.org/10.1242/jcs.261720</a>","ieee":"N. Gnyliukh <i>et al.</i>, “Role of dynamin-related proteins 2 and SH3P2 in clathrin-mediated endocytosis in Arabidopsis thaliana,” <i>Journal of Cell Science</i>, vol. 137, no. 8. The Company of Biologists, 2024.","ama":"Gnyliukh N, Johnson AJ, Nagel M, et al. Role of dynamin-related proteins 2 and SH3P2 in clathrin-mediated endocytosis in Arabidopsis thaliana. <i>Journal of Cell Science</i>. 2024;137(8). doi:<a href=\"https://doi.org/10.1242/jcs.261720\">10.1242/jcs.261720</a>","chicago":"Gnyliukh, Nataliia, Alexander J Johnson, MK Nagel, Aline Monzer, David Babic, Annamaria Hlavata, SS Alotaibi, E Isono, Martin Loose, and Jiří Friml. “Role of Dynamin-Related Proteins 2 and SH3P2 in Clathrin-Mediated Endocytosis in Arabidopsis Thaliana.” <i>Journal of Cell Science</i>. The Company of Biologists, 2024. <a href=\"https://doi.org/10.1242/jcs.261720\">https://doi.org/10.1242/jcs.261720</a>."},"acknowledged_ssus":[{"_id":"EM-Fac"},{"_id":"LifeSc"},{"_id":"Bio"}],"article_processing_charge":"Yes (via OA deal)","ec_funded":1,"tmp":{"legal_code_url":"https://creativecommons.org/licenses/by/4.0/legalcode","name":"Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)","image":"/images/cc_by.png","short":"CC BY (4.0)"},"OA_place":"publisher","month":"04","intvolume":"       137","publisher":"The Company of Biologists"},{"_id":"14510","corr_author":"1","status":"public","department":[{"_id":"GradSch"},{"_id":"JiFr"},{"_id":"MaLo"}],"file_date_updated":"2024-11-23T23:30:38Z","ddc":["570"],"degree_awarded":"PhD","related_material":{"record":[{"relation":"part_of_dissertation","status":"public","id":"14591"},{"relation":"part_of_dissertation","status":"public","id":"9887"},{"id":"8139","relation":"part_of_dissertation","status":"public"}]},"oa":1,"date_updated":"2026-06-21T22:30:42Z","month":"11","OA_place":"publisher","supervisor":[{"last_name":"Friml","full_name":"Friml, Jiří","id":"4159519E-F248-11E8-B48F-1D18A9856A87","first_name":"Jiří","orcid":"0000-0002-8302-7596"},{"first_name":"Martin","id":"462D4284-F248-11E8-B48F-1D18A9856A87","full_name":"Loose, Martin","last_name":"Loose","orcid":"0000-0001-7309-9724"}],"publisher":"Institute of Science and Technology Austria","citation":{"ieee":"N. Gnyliukh, “Mechanism of clathrin-coated vesicle  formation during endocytosis in plants,” Institute of Science and Technology Austria, 2023.","chicago":"Gnyliukh, Nataliia. “Mechanism of Clathrin-Coated Vesicle  Formation during Endocytosis in Plants.” Institute of Science and Technology Austria, 2023. <a href=\"https://doi.org/10.15479/at:ista:14510\">https://doi.org/10.15479/at:ista:14510</a>.","ama":"Gnyliukh N. Mechanism of clathrin-coated vesicle  formation during endocytosis in plants. 2023. doi:<a href=\"https://doi.org/10.15479/at:ista:14510\">10.15479/at:ista:14510</a>","short":"N. Gnyliukh, Mechanism of Clathrin-Coated Vesicle  Formation during Endocytosis in Plants, Institute of Science and Technology Austria, 2023.","ista":"Gnyliukh N. 2023. Mechanism of clathrin-coated vesicle  formation during endocytosis in plants. Institute of Science and Technology Austria.","apa":"Gnyliukh, N. (2023). <i>Mechanism of clathrin-coated vesicle  formation during endocytosis in plants</i>. Institute of Science and Technology Austria. <a href=\"https://doi.org/10.15479/at:ista:14510\">https://doi.org/10.15479/at:ista:14510</a>","mla":"Gnyliukh, Nataliia. <i>Mechanism of Clathrin-Coated Vesicle  Formation during Endocytosis in Plants</i>. Institute of Science and Technology Austria, 2023, doi:<a href=\"https://doi.org/10.15479/at:ista:14510\">10.15479/at:ista:14510</a>."},"has_accepted_license":"1","tmp":{"legal_code_url":"https://creativecommons.org/licenses/by/4.0/legalcode","name":"Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)","image":"/images/cc_by.png","short":"CC BY (4.0)"},"acknowledged_ssus":[{"_id":"EM-Fac"},{"_id":"Bio"},{"_id":"LifeSc"}],"ec_funded":1,"article_processing_charge":"No","page":"180","publication_status":"published","language":[{"iso":"eng"}],"type":"dissertation","title":"Mechanism of clathrin-coated vesicle  formation during endocytosis in plants","project":[{"call_identifier":"H2020","grant_number":"665385","name":"International IST Doctoral Program","_id":"2564DBCA-B435-11E9-9278-68D0E5697425"}],"alternative_title":["ISTA Thesis"],"date_created":"2023-11-10T09:10:06Z","user_id":"ba8df636-2132-11f1-aed0-ed93e2281fdd","publication_identifier":{"issn":["2663-337X"],"isbn":["978-3-99078-037-4"]},"year":"2023","doi":"10.15479/at:ista:14510","keyword":["Clathrin-Mediated Endocytosis","vesicle scission","Dynamin-Related Protein 2","SH3P2","TPLATE complex","Total internal reflection fluorescence microscopy","Arabidopsis thaliana"],"file":[{"file_size":20824903,"date_created":"2023-11-20T09:18:51Z","content_type":"application/vnd.openxmlformats-officedocument.wordprocessingml.document","date_updated":"2024-11-23T23:30:38Z","creator":"ngnyliuk","relation":"source_file","access_level":"closed","file_name":"Thesis_Gnyliukh_final_08_11_23.docx","file_id":"14567","checksum":"3d5e680bfc61f98e308c434f45cc9bd6","embargo_to":"open_access"},{"file_id":"14568","file_name":"Thesis_Gnyliukh_final_20_11_23.pdf","relation":"main_file","embargo":"2024-11-23","access_level":"open_access","checksum":"bfc96d47fc4e7e857dd71656097214a4","date_created":"2023-11-20T09:23:11Z","content_type":"application/pdf","file_size":24871844,"creator":"ngnyliuk","date_updated":"2024-11-23T23:30:38Z"}],"date_published":"2023-11-10T00:00:00Z","abstract":[{"lang":"eng","text":"Clathrin-mediated endocytosis (CME) is vital for the regulation of plant growth and\r\ndevelopment by controlling plasma membrane protein composition and cargo uptake. CME\r\nrelies on the precise recruitment control of protein regulators for vesicle maturation and\r\nrelease. During the early stages of endocytosis, an area of flat membrane is remodelled by\r\nproteins to create a spherical vesicle against intracellular forces. After the Clathrin-coated\r\nvesicle (CCV) is fully formed, scission machinery releases it from the plasma membrane,\r\nand cargo proceeds for recycling or degradation through early endosomes / Trans Golgi\r\nnetwork. Protein machineries that mediate membrane bending and vesicle release in plants\r\nare unknown. However, studies show, that plant endocytosis is actin independent, thus\r\nindicating that plants utilize a unique mechanism to mediate membrane bending against highturgor pressure compared to other model systems. First, by using biochemical and advanced\r\nlive microscopy approaches we investigate the TPLATE complex, a plant-specific\r\nendocytosis protein complex. We found that TPLATE is peripherally associated with\r\nclathrin-coated vesicles and localises at the rim of endocytosis events. Next, our study of\r\nplant Dynamin-related protein 1C (DRP1C), which was hypothesised previously to play a\r\nrole in vesicle release, shows the recruitment of the protein already at the early stages of\r\nendocytosis. Moreover, DRP1C assembles into organised ring-like structures and is able to\r\ninduce membrane deformation and tubulation, suggesting its role also in membrane bending\r\nduring early CME. Based on the data from mammalian and yeast systems, plant DynaminRelated Proteins 2 and SH3P2 protein are strong candidates to be part of the plant vesicle\r\nscission machinery; however, their precise role in plant CME has not been yet elucidated.\r\nHere, we characterised DRP2s and SH3P2 roles in CME by combining high-resolution\r\nimaging of endocytic events in vivo and protein characterisation. Although DRP2s and\r\nSH3P2 arrive together during late CME and physically interact, genetic analysis using\r\n∆sh3p1,2,3 mutant and complementation with non-DRP2-interacting SH3P2 variants suggest\r\nthat SH3P2 does not directly recruit DRP2s to the site of endocytosis. Summarising our\r\nresearch, these observations provide new important insights into the mechanism of plant\r\nCME and show that, despite plants posses many homologues of mammalian and yeast CME\r\ncomponents, they do not necessarily act in the same manner. "}],"oa_version":"Published Version","day":"10","author":[{"orcid":"0000-0002-2198-0509","full_name":"Gnyliukh, Nataliia","last_name":"Gnyliukh","first_name":"Nataliia","id":"390C1120-F248-11E8-B48F-1D18A9856A87"}]},{"author":[{"orcid":"0000-0002-2198-0509","first_name":"Nataliia","id":"390C1120-F248-11E8-B48F-1D18A9856A87","full_name":"Gnyliukh, Nataliia","last_name":"Gnyliukh"},{"id":"46A62C3A-F248-11E8-B48F-1D18A9856A87","first_name":"Alexander J","last_name":"Johnson","full_name":"Johnson, Alexander J","orcid":"0000-0002-2739-8843"},{"first_name":"Marie-Kristin","last_name":"Nagel","full_name":"Nagel, Marie-Kristin"},{"first_name":"Aline","id":"2DB5D88C-D7B3-11E9-B8FD-7907E6697425","full_name":"Monzer, Aline","last_name":"Monzer"},{"first_name":"Annamaria","id":"36062FEC-F248-11E8-B48F-1D18A9856A87","full_name":"Hlavata, Annamaria","last_name":"Hlavata"},{"first_name":"Erika","full_name":"Isono, Erika","last_name":"Isono"},{"first_name":"Martin","id":"462D4284-F248-11E8-B48F-1D18A9856A87","full_name":"Loose, Martin","last_name":"Loose","orcid":"0000-0001-7309-9724"},{"orcid":"0000-0002-8302-7596","first_name":"Jiří","id":"4159519E-F248-11E8-B48F-1D18A9856A87","full_name":"Friml, Jiří","last_name":"Friml"}],"acknowledged_ssus":[{"_id":"EM-Fac"},{"_id":"LifeSc"},{"_id":"Bio"}],"ec_funded":1,"article_processing_charge":"No","day":"10","oa_version":"Preprint","citation":{"short":"N. Gnyliukh, A.J. Johnson, M.-K. Nagel, A. Monzer, A. Hlavata, E. Isono, M. Loose, J. Friml, BioRxiv (n.d.).","ista":"Gnyliukh N, Johnson AJ, Nagel M-K, Monzer A, Hlavata A, Isono E, Loose M, Friml J. Role of dynamin-related proteins 2 and SH3P2 in clathrin-mediated endocytosis in plants. bioRxiv, <a href=\"https://doi.org/10.1101/2023.10.09.561523\">10.1101/2023.10.09.561523</a>.","mla":"Gnyliukh, Nataliia, et al. “Role of Dynamin-Related Proteins 2 and SH3P2 in Clathrin-Mediated Endocytosis in Plants.” <i>BioRxiv</i>, doi:<a href=\"https://doi.org/10.1101/2023.10.09.561523\">10.1101/2023.10.09.561523</a>.","apa":"Gnyliukh, N., Johnson, A. J., Nagel, M.-K., Monzer, A., Hlavata, A., Isono, E., … Friml, J. (n.d.). Role of dynamin-related proteins 2 and SH3P2 in clathrin-mediated endocytosis in plants. <i>bioRxiv</i>. <a href=\"https://doi.org/10.1101/2023.10.09.561523\">https://doi.org/10.1101/2023.10.09.561523</a>","ieee":"N. Gnyliukh <i>et al.</i>, “Role of dynamin-related proteins 2 and SH3P2 in clathrin-mediated endocytosis in plants,” <i>bioRxiv</i>. .","ama":"Gnyliukh N, Johnson AJ, Nagel M-K, et al. Role of dynamin-related proteins 2 and SH3P2 in clathrin-mediated endocytosis in plants. <i>bioRxiv</i>. doi:<a href=\"https://doi.org/10.1101/2023.10.09.561523\">10.1101/2023.10.09.561523</a>","chicago":"Gnyliukh, Nataliia, Alexander J Johnson, Marie-Kristin Nagel, Aline Monzer, Annamaria Hlavata, Erika Isono, Martin Loose, and Jiří Friml. “Role of Dynamin-Related Proteins 2 and SH3P2 in Clathrin-Mediated Endocytosis in Plants.” <i>BioRxiv</i>, n.d. <a href=\"https://doi.org/10.1101/2023.10.09.561523\">https://doi.org/10.1101/2023.10.09.561523</a>."},"main_file_link":[{"open_access":"1","url":"https://doi.org/10.1101/2023.10.09.561523"}],"abstract":[{"text":"Clathrin-mediated endocytosis (CME) is vital for the regulation of plant growth and development by controlling plasma membrane protein composition and cargo uptake. CME relies on the precise recruitment of regulators for vesicle maturation and release. Homologues of components of mammalian vesicle scission are strong candidates to be part of the scissin machinery in plants, but the precise roles of these proteins in this process is not fully understood. Here, we characterised the roles of Plant Dynamin-Related Proteins 2 (DRP2s) and SH3-domain containing protein 2 (SH3P2), the plant homologue to Dynamins’ recruiters, like Endophilin and Amphiphysin, in the CME by combining high-resolution imaging of endocytic events in vivo and characterisation of the purified proteins in vitro. Although DRP2s and SH3P2 arrive similarly late during CME and physically interact, genetic analysis of the Dsh3p1,2,3 triple-mutant and complementation assays with non-SH3P2-interacting DRP2 variants suggests that SH3P2 does not directly recruit DRP2s to the site of endocytosis. These observations imply that despite the presence of many well-conserved endocytic components, plants have acquired a distinct mechanism for CME. One Sentence Summary In contrast to predictions based on mammalian systems, plant Dynamin-related proteins 2 are recruited to the site of Clathrin-mediated endocytosis independently of BAR-SH3 proteins.","lang":"eng"}],"month":"10","publication":"bioRxiv","OA_place":"repository","date_published":"2023-10-10T00:00:00Z","doi":"10.1101/2023.10.09.561523","date_updated":"2026-06-21T22:30:40Z","year":"2023","user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","related_material":{"record":[{"status":"public","relation":"later_version","id":"15330"},{"status":"public","relation":"dissertation_contains","id":"14510"}]},"date_created":"2023-11-22T10:17:49Z","oa":1,"title":"Role of dynamin-related proteins 2 and SH3P2 in clathrin-mediated endocytosis in plants","type":"preprint","project":[{"call_identifier":"H2020","grant_number":"665385","name":"International IST Doctoral Program","_id":"2564DBCA-B435-11E9-9278-68D0E5697425"}],"department":[{"_id":"JiFr"},{"_id":"MaLo"},{"_id":"CaBe"}],"language":[{"iso":"eng"}],"publication_status":"draft","status":"public","_id":"14591","corr_author":"1"},{"acknowledgement":"We gratefully thank Julie Neveu and Dr. Amanda Barranco of the Grégory Vert laboratory for help preparing plants in France, Dr. Zuzana Gelova for help and advice with protoplast generation, Dr. Stéphane Vassilopoulos and Dr. Florian Schur for advice regarding EM tomography, Alejandro Marquiegui Alvaro for help with material generation, and Dr. Lukasz Kowalski for generously gifting us the mWasabi protein. This research was supported by the Scientific Service Units of Institute of Science and Technology Austria (IST Austria) through resources provided by the Electron Microscopy Facility, Lab Support Facility (particularly Dorota Jaworska), and the Bioimaging Facility. We acknowledge the Advanced Microscopy Facility of the Vienna BioCenter Core Facilities for use of the 3D SIM. For the mass spectrometry analysis of proteins, we acknowledge the University of Natural Resources and Life Sciences (BOKU) Core Facility Mass Spectrometry. This work was supported by the following funds: A.J. is supported by funding from the Austrian Science Fund I3630B25 to J.F. P.M. and E.B. are supported by Agence Nationale de la Recherche ANR-11-EQPX-0029 Morphoscope2 and ANR-10-INBS-04 France BioImaging. S.Y.B. is supported by the NSF No. 1121998 and 1614915. J.W. and D.V.D. are supported by the European Research Council Grant 682436 (to D.V.D.), a China Scholarship Council Grant 201508440249 (to J.W.), and by a Ghent University Special Research Co-funding Grant ST01511051 (to J.W.).","doi":"10.1073/pnas.2113046118","article_number":"e2113046118","year":"2021","publication_identifier":{"eissn":["1091-6490"]},"user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","date_created":"2021-08-11T14:11:43Z","type":"journal_article","title":"The TPLATE complex mediates membrane bending during plant clathrin-mediated endocytosis","project":[{"_id":"26538374-B435-11E9-9278-68D0E5697425","grant_number":"I03630","name":"Molecular mechanisms of endocytic cargo recognition in plants","call_identifier":"FWF"}],"isi":1,"language":[{"iso":"eng"}],"publication_status":"published","article_type":"original","author":[{"last_name":"Johnson","full_name":"Johnson, Alexander J","id":"46A62C3A-F248-11E8-B48F-1D18A9856A87","first_name":"Alexander J","orcid":"0000-0002-2739-8843"},{"first_name":"Dana A","last_name":"Dahhan","full_name":"Dahhan, Dana A"},{"orcid":"0000-0002-2198-0509","full_name":"Gnyliukh, Nataliia","last_name":"Gnyliukh","first_name":"Nataliia","id":"390C1120-F248-11E8-B48F-1D18A9856A87"},{"full_name":"Kaufmann, Walter","last_name":"Kaufmann","first_name":"Walter","id":"3F99E422-F248-11E8-B48F-1D18A9856A87","orcid":"0000-0001-9735-5315"},{"last_name":"Zheden","full_name":"Zheden, Vanessa","id":"39C5A68A-F248-11E8-B48F-1D18A9856A87","first_name":"Vanessa","orcid":"0000-0002-9438-4783"},{"orcid":"0000-0001-9732-3815","last_name":"Costanzo","full_name":"Costanzo, Tommaso","id":"D93824F4-D9BA-11E9-BB12-F207E6697425","first_name":"Tommaso"},{"first_name":"Pierre","last_name":"Mahou","full_name":"Mahou, Pierre"},{"full_name":"Hrtyan, Mónika","last_name":"Hrtyan","first_name":"Mónika","id":"45A71A74-F248-11E8-B48F-1D18A9856A87"},{"first_name":"Jie","full_name":"Wang, Jie","last_name":"Wang"},{"orcid":"0000-0002-2862-8372","id":"2A67C376-F248-11E8-B48F-1D18A9856A87","first_name":"Juan L","last_name":"Aguilera Servin","full_name":"Aguilera Servin, Juan L"},{"last_name":"van Damme","full_name":"van Damme, Daniël","first_name":"Daniël"},{"first_name":"Emmanuel","full_name":"Beaurepaire, Emmanuel","last_name":"Beaurepaire"},{"id":"462D4284-F248-11E8-B48F-1D18A9856A87","first_name":"Martin","last_name":"Loose","full_name":"Loose, Martin","orcid":"0000-0001-7309-9724"},{"last_name":"Bednarek","full_name":"Bednarek, Sebastian Y","first_name":"Sebastian Y"},{"orcid":"0000-0002-8302-7596","last_name":"Friml","full_name":"Friml, Jiří","id":"4159519E-F248-11E8-B48F-1D18A9856A87","first_name":"Jiří"}],"day":"14","pmid":1,"oa_version":"Published Version","issue":"51","abstract":[{"text":"Clathrin-mediated endocytosis is the major route of entry of cargos into cells and thus underpins many physiological processes. During endocytosis, an area of flat membrane is remodeled by proteins to create a spherical vesicle against intracellular forces. The protein machinery which mediates this membrane bending in plants is unknown. However, it is known that plant endocytosis is actin independent, thus indicating that plants utilize a unique mechanism to mediate membrane bending against high-turgor pressure compared to other model systems. Here, we investigate the TPLATE complex, a plant-specific endocytosis protein complex. It has been thought to function as a classical adaptor functioning underneath the clathrin coat. However, by using biochemical and advanced live microscopy approaches, we found that TPLATE is peripherally associated with clathrin-coated vesicles and localizes at the rim of endocytosis events. As this localization is more fitting to the protein machinery involved in membrane bending during endocytosis, we examined cells in which the TPLATE complex was disrupted and found that the clathrin structures present as flat patches. This suggests a requirement of the TPLATE complex for membrane bending during plant clathrin–mediated endocytosis. Next, we used in vitro biophysical assays to confirm that the TPLATE complex possesses protein domains with intrinsic membrane remodeling activity. These results redefine the role of the TPLATE complex and implicate it as a key component of the evolutionarily distinct plant endocytosis mechanism, which mediates endocytic membrane bending against the high-turgor pressure in plant cells.","lang":"eng"}],"publication":"Proceedings of the National Academy of Sciences of the United States of America","file":[{"file_size":2757340,"success":1,"date_created":"2021-12-15T08:59:40Z","content_type":"application/pdf","date_updated":"2021-12-15T08:59:40Z","creator":"cchlebak","access_level":"open_access","relation":"main_file","file_name":"2021_PNAS_Johnson.pdf","file_id":"10546","checksum":"8d01e72e22c4fb1584e72d8601947069"}],"external_id":{"isi":["000736417600043"],"pmid":["34907016"]},"date_published":"2021-12-14T00:00:00Z","date_updated":"2026-06-21T22:30:40Z","related_material":{"link":[{"relation":"earlier_version","url":"https://doi.org/10.1101/2021.04.26.441441"}],"record":[{"id":"14988","status":"public","relation":"research_data"},{"status":"public","relation":"dissertation_contains","id":"14510"}]},"oa":1,"scopus_import":"1","file_date_updated":"2021-12-15T08:59:40Z","department":[{"_id":"JiFr"},{"_id":"MaLo"},{"_id":"EvBe"},{"_id":"EM-Fac"},{"_id":"NanoFab"}],"ddc":["580"],"status":"public","corr_author":"1","_id":"9887","tmp":{"legal_code_url":"https://creativecommons.org/licenses/by/4.0/legalcode","name":"Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)","image":"/images/cc_by.png","short":"CC BY (4.0)"},"article_processing_charge":"No","acknowledged_ssus":[{"_id":"EM-Fac"},{"_id":"LifeSc"},{"_id":"Bio"}],"quality_controlled":"1","volume":118,"citation":{"ieee":"A. J. Johnson <i>et al.</i>, “The TPLATE complex mediates membrane bending during plant clathrin-mediated endocytosis,” <i>Proceedings of the National Academy of Sciences of the United States of America</i>, vol. 118, no. 51. National Academy of Sciences, 2021.","chicago":"Johnson, Alexander J, Dana A Dahhan, Nataliia Gnyliukh, Walter Kaufmann, Vanessa Zheden, Tommaso Costanzo, Pierre Mahou, et al. “The TPLATE Complex Mediates Membrane Bending during Plant Clathrin-Mediated Endocytosis.” <i>Proceedings of the National Academy of Sciences of the United States of America</i>. National Academy of Sciences, 2021. <a href=\"https://doi.org/10.1073/pnas.2113046118\">https://doi.org/10.1073/pnas.2113046118</a>.","ama":"Johnson AJ, Dahhan DA, Gnyliukh N, et al. The TPLATE complex mediates membrane bending during plant clathrin-mediated endocytosis. <i>Proceedings of the National Academy of Sciences of the United States of America</i>. 2021;118(51). doi:<a href=\"https://doi.org/10.1073/pnas.2113046118\">10.1073/pnas.2113046118</a>","short":"A.J. Johnson, D.A. Dahhan, N. Gnyliukh, W. Kaufmann, V. Zheden, T. Costanzo, P. Mahou, M. Hrtyan, J. Wang, J.L. Aguilera Servin, D. van Damme, E. Beaurepaire, M. Loose, S.Y. Bednarek, J. Friml, Proceedings of the National Academy of Sciences of the United States of America 118 (2021).","ista":"Johnson AJ, Dahhan DA, Gnyliukh N, Kaufmann W, Zheden V, Costanzo T, Mahou P, Hrtyan M, Wang J, Aguilera Servin JL, van Damme D, Beaurepaire E, Loose M, Bednarek SY, Friml J. 2021. The TPLATE complex mediates membrane bending during plant clathrin-mediated endocytosis. Proceedings of the National Academy of Sciences of the United States of America. 118(51), e2113046118.","apa":"Johnson, A. J., Dahhan, D. A., Gnyliukh, N., Kaufmann, W., Zheden, V., Costanzo, T., … Friml, J. (2021). The TPLATE complex mediates membrane bending during plant clathrin-mediated endocytosis. <i>Proceedings of the National Academy of Sciences of the United States of America</i>. National Academy of Sciences. <a href=\"https://doi.org/10.1073/pnas.2113046118\">https://doi.org/10.1073/pnas.2113046118</a>","mla":"Johnson, Alexander J., et al. “The TPLATE Complex Mediates Membrane Bending during Plant Clathrin-Mediated Endocytosis.” <i>Proceedings of the National Academy of Sciences of the United States of America</i>, vol. 118, no. 51, e2113046118, National Academy of Sciences, 2021, doi:<a href=\"https://doi.org/10.1073/pnas.2113046118\">10.1073/pnas.2113046118</a>."},"has_accepted_license":"1","intvolume":"       118","publisher":"National Academy of Sciences","month":"12"},{"has_accepted_license":"1","citation":{"short":"A.J. Johnson, N. Gnyliukh, W. Kaufmann, M. Narasimhan, G. Vert, S. Bednarek, J. Friml, Journal of Cell Science 133 (2020).","ista":"Johnson AJ, Gnyliukh N, Kaufmann W, Narasimhan M, Vert G, Bednarek S, Friml J. 2020. Experimental toolbox for quantitative evaluation of clathrin-mediated endocytosis in the plant model Arabidopsis. Journal of Cell Science. 133(15), jcs248062.","mla":"Johnson, Alexander J., et al. “Experimental Toolbox for Quantitative Evaluation of Clathrin-Mediated Endocytosis in the Plant Model Arabidopsis.” <i>Journal of Cell Science</i>, vol. 133, no. 15, jcs248062, The Company of Biologists, 2020, doi:<a href=\"https://doi.org/10.1242/jcs.248062\">10.1242/jcs.248062</a>.","apa":"Johnson, A. J., Gnyliukh, N., Kaufmann, W., Narasimhan, M., Vert, G., Bednarek, S., &#38; Friml, J. (2020). Experimental toolbox for quantitative evaluation of clathrin-mediated endocytosis in the plant model Arabidopsis. <i>Journal of Cell Science</i>. The Company of Biologists. <a href=\"https://doi.org/10.1242/jcs.248062\">https://doi.org/10.1242/jcs.248062</a>","ieee":"A. J. Johnson <i>et al.</i>, “Experimental toolbox for quantitative evaluation of clathrin-mediated endocytosis in the plant model Arabidopsis,” <i>Journal of Cell Science</i>, vol. 133, no. 15. The Company of Biologists, 2020.","chicago":"Johnson, Alexander J, Nataliia Gnyliukh, Walter Kaufmann, Madhumitha Narasimhan, G Vert, SY Bednarek, and Jiří Friml. “Experimental Toolbox for Quantitative Evaluation of Clathrin-Mediated Endocytosis in the Plant Model Arabidopsis.” <i>Journal of Cell Science</i>. The Company of Biologists, 2020. <a href=\"https://doi.org/10.1242/jcs.248062\">https://doi.org/10.1242/jcs.248062</a>.","ama":"Johnson AJ, Gnyliukh N, Kaufmann W, et al. Experimental toolbox for quantitative evaluation of clathrin-mediated endocytosis in the plant model Arabidopsis. <i>Journal of Cell Science</i>. 2020;133(15). doi:<a href=\"https://doi.org/10.1242/jcs.248062\">10.1242/jcs.248062</a>"},"quality_controlled":"1","volume":133,"acknowledged_ssus":[{"_id":"EM-Fac"},{"_id":"Bio"}],"article_processing_charge":"No","ec_funded":1,"month":"08","publisher":"The Company of Biologists","intvolume":"       133","oa":1,"related_material":{"record":[{"id":"14510","status":"public","relation":"dissertation_contains"}]},"date_updated":"2026-06-21T22:30:41Z","_id":"8139","status":"public","ddc":["575"],"file_date_updated":"2021-08-08T22:30:03Z","department":[{"_id":"JiFr"},{"_id":"EM-Fac"}],"scopus_import":"1","issue":"15","pmid":1,"oa_version":"Published Version","day":"06","author":[{"full_name":"Johnson, Alexander J","last_name":"Johnson","first_name":"Alexander J","id":"46A62C3A-F248-11E8-B48F-1D18A9856A87","orcid":"0000-0002-2739-8843"},{"orcid":"0000-0002-2198-0509","first_name":"Nataliia","id":"390C1120-F248-11E8-B48F-1D18A9856A87","full_name":"Gnyliukh, Nataliia","last_name":"Gnyliukh"},{"full_name":"Kaufmann, Walter","last_name":"Kaufmann","first_name":"Walter","id":"3F99E422-F248-11E8-B48F-1D18A9856A87","orcid":"0000-0001-9735-5315"},{"id":"44BF24D0-F248-11E8-B48F-1D18A9856A87","first_name":"Madhumitha","last_name":"Narasimhan","full_name":"Narasimhan, Madhumitha","orcid":"0000-0002-8600-0671"},{"full_name":"Vert, G","last_name":"Vert","first_name":"G"},{"last_name":"Bednarek","full_name":"Bednarek, SY","first_name":"SY"},{"orcid":"0000-0002-8302-7596","first_name":"Jiří","id":"4159519E-F248-11E8-B48F-1D18A9856A87","full_name":"Friml, Jiří","last_name":"Friml"}],"file":[{"file_size":15150403,"content_type":"application/pdf","date_created":"2020-11-26T17:12:51Z","date_updated":"2021-08-08T22:30:03Z","creator":"ajohnson","relation":"main_file","embargo":"2021-08-07","access_level":"open_access","file_id":"8815","file_name":"2020 - Johnson - JSC - plant CME toolbox.pdf","checksum":"2d11f79a0b4e0a380fb002b933da331a"}],"date_published":"2020-08-06T00:00:00Z","external_id":{"isi":["000561047900021"],"pmid":["32616560"]},"publication":"Journal of Cell Science","abstract":[{"lang":"eng","text":"Clathrin-mediated endocytosis (CME) is a crucial cellular process implicated in many aspects of plant growth, development, intra- and inter-cellular signaling, nutrient uptake and pathogen defense. Despite these significant roles, little is known about the precise molecular details of how it functions in planta. In order to facilitate the direct quantitative study of plant CME, here we review current routinely used methods and present refined, standardized quantitative imaging protocols which allow the detailed characterization of CME at multiple scales in plant tissues. These include: (i) an efficient electron microscopy protocol for the imaging of Arabidopsis CME vesicles in situ, thus providing a method for the detailed characterization of the ultra-structure of clathrin-coated vesicles; (ii) a detailed protocol and analysis for quantitative live-cell fluorescence microscopy to precisely examine the temporal interplay of endocytosis components during single CME events; (iii) a semi-automated analysis to allow the quantitative characterization of global internalization of cargos in whole plant tissues; and (iv) an overview and validation of useful genetic and pharmacological tools to interrogate the molecular mechanisms and function of CME in intact plant samples."}],"date_created":"2020-07-21T08:58:19Z","user_id":"c635000d-4b10-11ee-a964-aac5a93f6ac1","year":"2020","article_number":"jcs248062","publication_identifier":{"eissn":["1477-9137"],"issn":["0021-9533"]},"doi":"10.1242/jcs.248062","acknowledgement":"This paper is dedicated to the memory of Christien Merrifield. He pioneered quantitative\r\nimaging approaches in mammalian CME and his mentorship inspired the development of all\r\nthe analysis methods presented here. His joy in research, pure scientific curiosity and\r\nmicroscopy excellence remain a constant inspiration. We thank Daniel Van Damme for gifting\r\nus the CLC2-GFP x TPLATE-TagRFP plants used in this manuscript. We further thank the\r\nScientific Service Units at IST Austria; specifically, the Electron Microscopy Facility for\r\ntechnical assistance (in particular Vanessa Zheden) and the BioImaging Facility BioImaging\r\nFacility for access to equipment. ","article_type":"original","publication_status":"published","language":[{"iso":"eng"}],"isi":1,"project":[{"call_identifier":"FWF","_id":"26538374-B435-11E9-9278-68D0E5697425","grant_number":"I03630","name":"Molecular mechanisms of endocytic cargo recognition in plants"},{"call_identifier":"H2020","name":"International IST Doctoral Program","grant_number":"665385","_id":"2564DBCA-B435-11E9-9278-68D0E5697425"}],"type":"journal_article","title":"Experimental toolbox for quantitative evaluation of clathrin-mediated endocytosis in the plant model Arabidopsis"}]