[{"author":[{"full_name":"Hlavata, Annamaria","last_name":"Hlavata","first_name":"Annamaria","id":"36062FEC-F248-11E8-B48F-1D18A9856A87"},{"full_name":"Neuditschko, Benjamin","last_name":"Neuditschko","first_name":"Benjamin"},{"first_name":"Ulla","last_name":"Schellhaas","full_name":"Schellhaas, Ulla"},{"first_name":"Clemens","last_name":"Plaschka","full_name":"Plaschka, Clemens"},{"full_name":"Herzog, Franz","last_name":"Herzog","first_name":"Franz"},{"orcid":"0000-0003-0893-7036","last_name":"Bernecky","id":"2CB9DFE2-F248-11E8-B48F-1D18A9856A87","first_name":"Carrie A","full_name":"Bernecky, Carrie A"}],"oa_version":"None","title":"Structure of cytoplasmic RNA polymerase II","date_created":"2025-12-11T13:33:27Z","date_published":"2025-12-10T00:00:00Z","abstract":[{"lang":"eng","text":"RNA polymerase II (Pol II) must be assembled in the cytoplasm before it enters the nucleus, where it transcribes protein-coding genes. Although transcription by Pol II is intensively studied, how this central multi-subunit enzyme is made and the role of dedicated factors remains unclear. Here, we report the integrative structural analysis of a native human Pol II from the cytoplasm captured near the end of biogenesis. The complex contained Gdown1 and three biogenesis factors – RPAP2 and the critical small GTPases GPN1 and GPN3. Cryo-EM analysis of the complex revealed how Gdown1 and RPAP2 associate with Pol II and prevent the premature association of transcription factors. Further biochemical and cryo-EM analysis revealed how RPAP2 recruits GPN1–GPN3 to the complex, and how the assembly of the RPAP2–GPN1–GPN3 complex is controlled by GTP hydrolysis. The combined results uncover a network of interactions that chaperone cytoplasmic Pol II to prevent aberrant interactions, reveal a GTP-controlled switch during the final stages of Pol II biogenesis, and suggest a general mechanism for the action of GPN-loop GTPase family of enzymes."}],"type":"preprint","article_processing_charge":"No","acknowledgement":"We thank A. Salmazo for assistance with Pol II purification. We thank staff at the VBCF Proteomics facility for immunoprecipitation-mass spectrometry analysis, and J.A. Stopp for assistance with IP-MS data visualization. This research was further supported by the Scientific Service Units (SSUs) of IST Austria through resources provided by the Lab Support Facility (LSF), Electron Microscopy (EMF), Scientific Computing (SciComp), and the Preclinical Facility (PCF).","year":"2025","oa":1,"main_file_link":[{"open_access":"1","url":"https://doi.org/10.64898/2025.12.10.692585"}],"language":[{"iso":"eng"}],"publication_status":"published","corr_author":"1","publisher":"bioRxiv","_id":"20804","status":"public","date_updated":"2025-12-15T09:48:22Z","citation":{"short":"A. Hlavata, B. Neuditschko, U. Schellhaas, C. Plaschka, F. Herzog, C. Bernecky, (2025).","apa":"Hlavata, A., Neuditschko, B., Schellhaas, U., Plaschka, C., Herzog, F., &#38; Bernecky, C. (2025). Structure of cytoplasmic RNA polymerase II. bioRxiv. <a href=\"https://doi.org/10.64898/2025.12.10.692585\">https://doi.org/10.64898/2025.12.10.692585</a>","ista":"Hlavata A, Neuditschko B, Schellhaas U, Plaschka C, Herzog F, Bernecky C. 2025. Structure of cytoplasmic RNA polymerase II. <a href=\"https://doi.org/10.64898/2025.12.10.692585\">10.64898/2025.12.10.692585</a>.","mla":"Hlavata, Annamaria, et al. <i>Structure of Cytoplasmic RNA Polymerase II</i>. bioRxiv, 2025, doi:<a href=\"https://doi.org/10.64898/2025.12.10.692585\">10.64898/2025.12.10.692585</a>.","ama":"Hlavata A, Neuditschko B, Schellhaas U, Plaschka C, Herzog F, Bernecky C. Structure of cytoplasmic RNA polymerase II. 2025. doi:<a href=\"https://doi.org/10.64898/2025.12.10.692585\">10.64898/2025.12.10.692585</a>","ieee":"A. Hlavata, B. Neuditschko, U. Schellhaas, C. Plaschka, F. Herzog, and C. Bernecky, “Structure of cytoplasmic RNA polymerase II.” bioRxiv, 2025.","chicago":"Hlavata, Annamaria, Benjamin Neuditschko, Ulla Schellhaas, Clemens Plaschka, Franz Herzog, and Carrie Bernecky. “Structure of Cytoplasmic RNA Polymerase II.” bioRxiv, 2025. <a href=\"https://doi.org/10.64898/2025.12.10.692585\">https://doi.org/10.64898/2025.12.10.692585</a>."},"month":"12","department":[{"_id":"CaBe"}],"user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","doi":"10.64898/2025.12.10.692585","acknowledged_ssus":[{"_id":"LifeSc"},{"_id":"EM-Fac"},{"_id":"ScienComp"},{"_id":"PreCl"}],"day":"10"},{"acknowledgement":"I would also like to acknowledge the ISTA Facilities: Lab Support Facility, Protein Services and Electron Microscopy Facility (EMF) and Scientific Computing. EMF for their support during data collections and troubleshooting, especially Valentin. Scientific Computing for solving quickly any issues related with cluster.","OA_place":"publisher","year":"2025","language":[{"iso":"eng"}],"corr_author":"1","publisher":"Institute of Science and Technology Austria","_id":"19431","page":"83","date_updated":"2026-04-07T11:46:32Z","citation":{"chicago":"Hlavata, Annamaria. “Regulation of Cytoplasmic RNA Polymerase II.” Institute of Science and Technology Austria, 2025. <a href=\"https://doi.org/10.15479/10.15479/AT-ISTA-19431\">https://doi.org/10.15479/10.15479/AT-ISTA-19431</a>.","mla":"Hlavata, Annamaria. <i>Regulation of Cytoplasmic RNA Polymerase II</i>. Institute of Science and Technology Austria, 2025, doi:<a href=\"https://doi.org/10.15479/10.15479/AT-ISTA-19431\">10.15479/10.15479/AT-ISTA-19431</a>.","ama":"Hlavata A. Regulation of Cytoplasmic RNA Polymerase II. 2025. doi:<a href=\"https://doi.org/10.15479/10.15479/AT-ISTA-19431\">10.15479/10.15479/AT-ISTA-19431</a>","ieee":"A. Hlavata, “Regulation of Cytoplasmic RNA Polymerase II,” Institute of Science and Technology Austria, 2025.","short":"A. Hlavata, Regulation of Cytoplasmic RNA Polymerase II, Institute of Science and Technology Austria, 2025.","apa":"Hlavata, A. (2025). <i>Regulation of Cytoplasmic RNA Polymerase II</i>. Institute of Science and Technology Austria. <a href=\"https://doi.org/10.15479/10.15479/AT-ISTA-19431\">https://doi.org/10.15479/10.15479/AT-ISTA-19431</a>","ista":"Hlavata A. 2025. Regulation of Cytoplasmic RNA Polymerase II. Institute of Science and Technology Austria."},"month":"03","file":[{"file_size":23506747,"checksum":"b7ddf424ffe95f8c767c53c8bb62d4f3","date_created":"2025-03-24T12:48:36Z","file_name":"PhD_Thesis_Hlavata_final_submission.docx","creator":"ahlavata","file_id":"19448","relation":"source_file","content_type":"application/vnd.openxmlformats-officedocument.wordprocessingml.document","embargo_to":"open_access","access_level":"closed","date_updated":"2026-03-20T23:30:04Z"},{"relation":"main_file","content_type":"application/pdf","embargo":"2026-03-20","date_updated":"2026-03-20T23:30:04Z","access_level":"open_access","date_created":"2025-03-24T12:51:10Z","checksum":"6c5a59c9bac467c3d0b3ffb8ea6d9fd4","file_size":9478591,"file_name":"PhD_Thesis_Hlavata_final_submission_update.pdf","creator":"ahlavata","file_id":"19449"}],"author":[{"first_name":"Annamaria","id":"36062FEC-F248-11E8-B48F-1D18A9856A87","last_name":"Hlavata","full_name":"Hlavata, Annamaria"}],"title":"Regulation of Cytoplasmic RNA Polymerase II","alternative_title":["ISTA Thesis"],"date_published":"2025-03-20T00:00:00Z","has_accepted_license":"1","type":"dissertation","ddc":["572"],"article_processing_charge":"No","publication_identifier":{"eissn":["2663-337X"],"isbn":["978-3-99078-055-8"]},"oa":1,"file_date_updated":"2026-03-20T23:30:04Z","publication_status":"published","status":"public","department":[{"_id":"GradSch"},{"_id":"CaBe"}],"user_id":"ba8df636-2132-11f1-aed0-ed93e2281fdd","acknowledged_ssus":[{"_id":"LifeSc"},{"_id":"EM-Fac"},{"_id":"ScienComp"}],"doi":"10.15479/10.15479/AT-ISTA-19431","day":"20","oa_version":"Published Version","supervisor":[{"orcid":"0000-0003-0893-7036","full_name":"Bernecky, Carrie A","id":"2CB9DFE2-F248-11E8-B48F-1D18A9856A87","first_name":"Carrie A","last_name":"Bernecky"}],"date_created":"2025-03-20T12:52:47Z","abstract":[{"lang":"eng","text":"Gene expression is crucial for cell differentiation, development and survival of\r\norganisms. It consists of several steps, starting with transcription that is mediated by\r\nRNA polymerases. These are protein machineries transcribing and producing different\r\ntypes of RNAs. Although, the individual steps of transcription by RNA polymerase II\r\n(Pol II) as well as the structure of Pol II has been extensively studied, surprisingly,\r\nthere is still little known about its regulation and assembly in cytoplasm. Among the\r\nproteins that are important in biogenesis of Pol II are RNA polymerase II associating\r\nproteins (RPAP) and small GPN-loop GTPases (GPN). Both of these protein groups\r\nwere shown to take essential part in assembly of Pol II.\r\nThe aim of this project was to deepen our knowledge in regulation of Pol II in\r\nthe cytoplasm as well as the proteins involved in this process. Techniques of structural\r\nbiology, biochemistry and cell biology were employed to study and characterize cytoplasmic Pol II and its interacting partners.\r\nThis study shows for the first time the structure of cytoplasmic Pol II at high\r\nresolution. The structure also reveals proteins interacting with Pol II in cytoplasm,\r\nnamely GDOWN1, RPAP2. Comparing the structure of cytoplasmic Pol II with transcribing Pol II revealed striking difference in clamp region that is not in closed state.\r\nFurthermore, GDOWN1 and RPAP2 make steric clashes with various transcription\r\nfactors bound to Pol II during different stages of transcription. Even though GPN1 and\r\nGPN3 proteins were not resolved in the cytoplasmic Pol II structure, they are part of\r\nthe complex and their interaction with Pol II was confirmed in vitro. RPAP2 stabilizes\r\nthese proteins on Pol II and several experiments suggest that they interact with the\r\nclamp region. In addition, GDOWN1, RPAP2 and GPNs might keep clamp in open or\r\npartially open state. Based on these results I propose a novel model of regulation of\r\nPol II in cytoplasm. GDOWN1, RPAP2, GPN1 and GPN3 bind to Pol II in cytoplasm\r\nand doing so they can prevent pre-mature binding of DNA or RNA and different transcription factors to Pol II in cytoplasm or before engaging in transcription nucleus.\r\nThis research contributes to the current knowledge of molecular mechanisms\r\nof Pol II regulation in cytoplasm."}],"degree_awarded":"PhD"},{"OA_type":"hybrid","pmid":1,"oa_version":"Published Version","volume":137,"date_created":"2024-04-19T09:54:59Z","article_type":"original","article_number":"jcs.261720","related_material":{"record":[{"status":"public","id":"14591","relation":"earlier_version"}]},"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."}],"scopus_import":"1","external_id":{"isi":["001266917100005"],"pmid":["38506228"]},"tmp":{"short":"CC BY (4.0)","image":"/images/cc_by.png","legal_code_url":"https://creativecommons.org/licenses/by/4.0/legalcode","name":"Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)"},"intvolume":"       137","issue":"8","file_date_updated":"2025-01-09T08:41:16Z","oa":1,"publication_status":"published","status":"public","acknowledged_ssus":[{"_id":"EM-Fac"},{"_id":"LifeSc"},{"_id":"Bio"}],"doi":"10.1242/jcs.261720","department":[{"_id":"MaLo"},{"_id":"JiFr"},{"_id":"CaBe"}],"user_id":"317138e5-6ab7-11ef-aa6d-ffef3953e345","project":[{"name":"International IST Doctoral Program","grant_number":"665385","_id":"2564DBCA-B435-11E9-9278-68D0E5697425","call_identifier":"H2020"},{"name":"Peptide receptors for auxin canalization in Arabidopsis","grant_number":"I06123","_id":"bd76d395-d553-11ed-ba76-f678c14f9033"}],"day":"01","isi":1,"ec_funded":1,"title":"Role of dynamin-related proteins 2 and SH3P2 in clathrin-mediated endocytosis in Arabidopsis thaliana","author":[{"full_name":"Gnyliukh, Nataliia","id":"390C1120-F248-11E8-B48F-1D18A9856A87","first_name":"Nataliia","last_name":"Gnyliukh","orcid":"0000-0002-2198-0509"},{"full_name":"Johnson, Alexander J","last_name":"Johnson","id":"46A62C3A-F248-11E8-B48F-1D18A9856A87","first_name":"Alexander J","orcid":"0000-0002-2739-8843"},{"last_name":"Nagel","first_name":"MK","full_name":"Nagel, MK"},{"first_name":"Aline","id":"2DB5D88C-D7B3-11E9-B8FD-7907E6697425","last_name":"Monzer","full_name":"Monzer, Aline"},{"full_name":"Babic, David","first_name":"David","id":"db566d23-f6e0-11ea-865d-e6f270e968e7","last_name":"Babic"},{"full_name":"Hlavata, Annamaria","last_name":"Hlavata","first_name":"Annamaria","id":"36062FEC-F248-11E8-B48F-1D18A9856A87"},{"full_name":"Alotaibi, SS","first_name":"SS","last_name":"Alotaibi"},{"full_name":"Isono, E","last_name":"Isono","first_name":"E"},{"orcid":"0000-0001-7309-9724","last_name":"Loose","id":"462D4284-F248-11E8-B48F-1D18A9856A87","first_name":"Martin","full_name":"Loose, Martin"},{"orcid":"0000-0002-8302-7596","id":"4159519E-F248-11E8-B48F-1D18A9856A87","first_name":"Jiří","last_name":"Friml","full_name":"Friml, Jiří"}],"date_published":"2024-04-01T00:00:00Z","type":"journal_article","has_accepted_license":"1","publication":"Journal of Cell Science","ddc":["570"],"article_processing_charge":"Yes (via OA deal)","publication_identifier":{"issn":["0021-9533"],"eissn":["1477-9137"]},"quality_controlled":"1","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).","OA_place":"publisher","year":"2024","corr_author":"1","publisher":"The Company of Biologists","language":[{"iso":"eng"}],"month":"04","_id":"15330","date_updated":"2025-09-04T13:49:45Z","citation":{"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>.","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>","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>.","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.","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>","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."},"file":[{"checksum":"6dc023f0cc7052ad3cf0a42589d2e30f","file_size":25845948,"date_created":"2025-01-09T08:41:16Z","file_name":"2024_JourCellScience_Gnyliukh.pdf","creator":"dernst","file_id":"18792","relation":"main_file","content_type":"application/pdf","success":1,"date_updated":"2025-01-09T08:41:16Z","access_level":"open_access"}]},{"corr_author":"1","publication_status":"draft","language":[{"iso":"eng"}],"month":"10","status":"public","_id":"14591","date_updated":"2026-06-18T22:30:30Z","citation":{"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>.","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>","ieee":"N. Gnyliukh <i>et al.</i>, “Role of dynamin-related proteins 2 and SH3P2 in clathrin-mediated endocytosis in plants,” <i>bioRxiv</i>. .","short":"N. Gnyliukh, A.J. Johnson, M.-K. Nagel, A. Monzer, A. Hlavata, E. Isono, M. Loose, J. Friml, BioRxiv (n.d.).","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>","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>.","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>."},"acknowledged_ssus":[{"_id":"EM-Fac"},{"_id":"LifeSc"},{"_id":"Bio"}],"doi":"10.1101/2023.10.09.561523","user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","department":[{"_id":"JiFr"},{"_id":"MaLo"},{"_id":"CaBe"}],"project":[{"name":"International IST Doctoral Program","call_identifier":"H2020","_id":"2564DBCA-B435-11E9-9278-68D0E5697425","grant_number":"665385"}],"day":"10","OA_place":"repository","year":"2023","oa":1,"main_file_link":[{"url":"https://doi.org/10.1101/2023.10.09.561523","open_access":"1"}],"publication":"bioRxiv","article_processing_charge":"No","ec_funded":1,"title":"Role of dynamin-related proteins 2 and SH3P2 in clathrin-mediated endocytosis in plants","oa_version":"Preprint","author":[{"id":"390C1120-F248-11E8-B48F-1D18A9856A87","first_name":"Nataliia","last_name":"Gnyliukh","full_name":"Gnyliukh, Nataliia","orcid":"0000-0002-2198-0509"},{"orcid":"0000-0002-2739-8843","full_name":"Johnson, Alexander J","id":"46A62C3A-F248-11E8-B48F-1D18A9856A87","first_name":"Alexander J","last_name":"Johnson"},{"first_name":"Marie-Kristin","last_name":"Nagel","full_name":"Nagel, Marie-Kristin"},{"first_name":"Aline","id":"2DB5D88C-D7B3-11E9-B8FD-7907E6697425","last_name":"Monzer","full_name":"Monzer, Aline"},{"full_name":"Hlavata, Annamaria","id":"36062FEC-F248-11E8-B48F-1D18A9856A87","first_name":"Annamaria","last_name":"Hlavata"},{"last_name":"Isono","first_name":"Erika","full_name":"Isono, Erika"},{"orcid":"0000-0001-7309-9724","last_name":"Loose","first_name":"Martin","id":"462D4284-F248-11E8-B48F-1D18A9856A87","full_name":"Loose, Martin"},{"full_name":"Friml, Jiří","last_name":"Friml","first_name":"Jiří","id":"4159519E-F248-11E8-B48F-1D18A9856A87","orcid":"0000-0002-8302-7596"}],"date_published":"2023-10-10T00:00:00Z","date_created":"2023-11-22T10:17:49Z","related_material":{"record":[{"status":"public","id":"15330","relation":"later_version"},{"id":"14510","relation":"dissertation_contains","status":"public"}]},"type":"preprint","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 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."}]}]