[{"page":"180","language":[{"iso":"eng"}],"department":[{"_id":"GradSch"},{"_id":"JiFr"},{"_id":"MaLo"}],"publication_status":"published","month":"11","license":"https://creativecommons.org/licenses/by/4.0/","date_created":"2023-11-10T09:10:06Z","article_processing_charge":"No","degree_awarded":"PhD","project":[{"_id":"2564DBCA-B435-11E9-9278-68D0E5697425","name":"International IST Doctoral Program","call_identifier":"H2020","grant_number":"665385"}],"related_material":{"record":[{"relation":"part_of_dissertation","id":"14591","status":"public"},{"status":"public","id":"9887","relation":"part_of_dissertation"},{"relation":"part_of_dissertation","id":"8139","status":"public"}]},"corr_author":"1","author":[{"full_name":"Gnyliukh, Nataliia","last_name":"Gnyliukh","id":"390C1120-F248-11E8-B48F-1D18A9856A87","orcid":"0000-0002-2198-0509","first_name":"Nataliia"}],"publication_identifier":{"issn":["2663-337X"],"isbn":["978-3-99078-037-4"]},"title":"Mechanism of clathrin-coated vesicle  formation during endocytosis in plants","oa":1,"doi":"10.15479/at:ista:14510","type":"dissertation","status":"public","_id":"14510","oa_version":"Published Version","day":"10","supervisor":[{"orcid":"0000-0002-8302-7596","first_name":"Jiří","full_name":"Friml, Jiří","last_name":"Friml","id":"4159519E-F248-11E8-B48F-1D18A9856A87"},{"first_name":"Martin","orcid":"0000-0001-7309-9724","id":"462D4284-F248-11E8-B48F-1D18A9856A87","last_name":"Loose","full_name":"Loose, Martin"}],"year":"2023","citation":{"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>.","short":"N. Gnyliukh, Mechanism of Clathrin-Coated Vesicle  Formation during Endocytosis in Plants, Institute of Science and Technology Austria, 2023.","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>.","ista":"Gnyliukh N. 2023. Mechanism of clathrin-coated vesicle  formation during endocytosis in plants. Institute of Science and Technology Austria.","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>","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>","ieee":"N. Gnyliukh, “Mechanism of clathrin-coated vesicle  formation during endocytosis in plants,” Institute of Science and Technology Austria, 2023."},"publisher":"Institute of Science and Technology Austria","acknowledged_ssus":[{"_id":"EM-Fac"},{"_id":"Bio"},{"_id":"LifeSc"}],"file":[{"relation":"source_file","access_level":"closed","checksum":"3d5e680bfc61f98e308c434f45cc9bd6","creator":"ngnyliuk","content_type":"application/vnd.openxmlformats-officedocument.wordprocessingml.document","date_updated":"2024-11-23T23:30:38Z","file_id":"14567","embargo_to":"open_access","date_created":"2023-11-20T09:18:51Z","file_size":20824903,"file_name":"Thesis_Gnyliukh_final_08_11_23.docx"},{"embargo":"2024-11-23","date_created":"2023-11-20T09:23:11Z","file_size":24871844,"file_name":"Thesis_Gnyliukh_final_20_11_23.pdf","file_id":"14568","creator":"ngnyliuk","content_type":"application/pdf","date_updated":"2024-11-23T23:30:38Z","relation":"main_file","access_level":"open_access","checksum":"bfc96d47fc4e7e857dd71656097214a4"}],"alternative_title":["ISTA Thesis"],"date_updated":"2026-10-26T23:30:39Z","user_id":"ba8df636-2132-11f1-aed0-ed93e2281fdd","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. "}],"file_date_updated":"2024-11-23T23:30:38Z","ec_funded":1,"ddc":["570"],"date_published":"2023-11-10T00:00:00Z","has_accepted_license":"1","fulldoi":"https://doi.org/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"],"tmp":{"image":"/images/cc_by.png","short":"CC BY (4.0)","legal_code_url":"https://creativecommons.org/licenses/by/4.0/legalcode","name":"Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)"},"OA_place":"publisher"},{"fulldoi":"https://doi.org/10.1038/s41541-022-00566-x","article_type":"original","tmp":{"image":"/images/cc_by.png","short":"CC BY (4.0)","legal_code_url":"https://creativecommons.org/licenses/by/4.0/legalcode","name":"Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)"},"keyword":["Pharmacology (medical)","Infectious Diseases","Pharmacology","Immunology","SARS-COV-2","COVID"],"ddc":["570"],"date_published":"2022-11-15T00:00:00Z","has_accepted_license":"1","abstract":[{"text":"Replication-incompetent adenoviral vectors have been extensively used as a platform for vaccine design, with at least four anti-COVID-19 vaccines authorized to date. These vaccines elicit neutralizing antibody responses directed against SARS-CoV-2 Spike protein and confer significant level of protection against SARS-CoV-2 infection. Immunization with adenovirus-vectored vaccines is known to be accompanied by the production of anti-vector antibodies, which may translate into reduced efficacy of booster or repeated rounds of revaccination. Here, we used blood samples from patients who received an adenovirus-based Gam-COVID-Vac vaccine to address the question of whether anti-vector antibodies may influence the magnitude of SARS-CoV-2-specific humoral response after booster vaccination. We observed that rAd26-based prime vaccination with Gam-COVID-Vac induced the development of Ad26-neutralizing antibodies, which persisted in circulation for at least 9 months. Our analysis further indicates that high pre-boost Ad26 neutralizing antibody titers do not appear to affect the humoral immunogenicity of the Gam-COVID-Vac boost. The titers of anti-SARS-CoV-2 RBD IgGs and antibodies, which neutralized both the wild type and the circulating variants of concern of SARS-CoV-2 such as Delta and Omicron, were independent of the pre-boost levels of Ad26-neutralizing antibodies. Thus, our results support the development of repeated immunization schedule with adenovirus-based COVID-19 vaccines.","lang":"eng"}],"file_date_updated":"2023-01-23T11:22:09Z","user_id":"4359f0d1-fa6c-11eb-b949-802e58b17ae8","date_updated":"2023-08-04T08:52:40Z","file":[{"file_id":"12347","success":1,"date_created":"2023-01-23T11:22:09Z","file_size":1856046,"file_name":"2022_njpVaccines_Byazrova.pdf","relation":"main_file","access_level":"open_access","checksum":"ddaac096381565b2b4b7dcc34cdbc4ee","creator":"dernst","date_updated":"2023-01-23T11:22:09Z","content_type":"application/pdf"}],"intvolume":"         7","external_id":{"isi":["000884278600004"],"pmid":["36379998"]},"acknowledgement":"We thank Sergey Kulemzin, Grigory Efimov, Yuri Lebedin, Alexander Taranin and Rudolf Valenta for providing reagents. Figures were created with the help of BioRender.com. This work was supported by the Russian Science Foundation (Project 21-15-00286). Byazrova M.G. was supported by the RUDN University Strategic Academic Leadership Program.","year":"2022","citation":{"ieee":"M. G. Byazrova <i>et al.</i>, “Anti-Ad26 humoral immunity does not compromise SARS-COV-2 neutralizing antibody responses following Gam-COVID-Vac booster vaccination,” <i>npj Vaccines</i>, vol. 7. Springer Nature, 2022.","apa":"Byazrova, M. G., Astakhova, E. A., Minnegalieva, A., Sukhova, M. M., Mikhailov, A. A., Prilipov, A. G., … Filatov, A. V. (2022). Anti-Ad26 humoral immunity does not compromise SARS-COV-2 neutralizing antibody responses following Gam-COVID-Vac booster vaccination. <i>Npj Vaccines</i>. Springer Nature. <a href=\"https://doi.org/10.1038/s41541-022-00566-x\">https://doi.org/10.1038/s41541-022-00566-x</a>","ista":"Byazrova MG, Astakhova EA, Minnegalieva A, Sukhova MM, Mikhailov AA, Prilipov AG, Gorchakov AA, Filatov AV. 2022. Anti-Ad26 humoral immunity does not compromise SARS-COV-2 neutralizing antibody responses following Gam-COVID-Vac booster vaccination. npj Vaccines. 7, 145.","ama":"Byazrova MG, Astakhova EA, Minnegalieva A, et al. Anti-Ad26 humoral immunity does not compromise SARS-COV-2 neutralizing antibody responses following Gam-COVID-Vac booster vaccination. <i>npj Vaccines</i>. 2022;7. doi:<a href=\"https://doi.org/10.1038/s41541-022-00566-x\">10.1038/s41541-022-00566-x</a>","mla":"Byazrova, Maria G., et al. “Anti-Ad26 Humoral Immunity Does Not Compromise SARS-COV-2 Neutralizing Antibody Responses Following Gam-COVID-Vac Booster Vaccination.” <i>Npj Vaccines</i>, vol. 7, 145, Springer Nature, 2022, doi:<a href=\"https://doi.org/10.1038/s41541-022-00566-x\">10.1038/s41541-022-00566-x</a>.","short":"M.G. Byazrova, E.A. Astakhova, A. Minnegalieva, M.M. Sukhova, A.A. Mikhailov, A.G. Prilipov, A.A. Gorchakov, A.V. Filatov, Npj Vaccines 7 (2022).","chicago":"Byazrova, Maria G., Ekaterina A. Astakhova, Aygul Minnegalieva, Maria M. Sukhova, Artem A. Mikhailov, Alexey G. Prilipov, Andrey A. Gorchakov, and Alexander V. Filatov. “Anti-Ad26 Humoral Immunity Does Not Compromise SARS-COV-2 Neutralizing Antibody Responses Following Gam-COVID-Vac Booster Vaccination.” <i>Npj Vaccines</i>. Springer Nature, 2022. <a href=\"https://doi.org/10.1038/s41541-022-00566-x\">https://doi.org/10.1038/s41541-022-00566-x</a>."},"publisher":"Springer Nature","day":"15","isi":1,"oa_version":"Published Version","_id":"12131","status":"public","article_number":"145","doi":"10.1038/s41541-022-00566-x","type":"journal_article","publication_identifier":{"issn":["2059-0105"]},"pmid":1,"oa":1,"title":"Anti-Ad26 humoral immunity does not compromise SARS-COV-2 neutralizing antibody responses following Gam-COVID-Vac booster vaccination","author":[{"last_name":"Byazrova","full_name":"Byazrova, Maria G.","first_name":"Maria G."},{"first_name":"Ekaterina A.","last_name":"Astakhova","full_name":"Astakhova, Ekaterina A."},{"id":"87DF77F0-1D9A-11EA-B6AE-CE443DDC885E","last_name":"Minnegalieva","full_name":"Minnegalieva, Aygul","first_name":"Aygul"},{"first_name":"Maria M.","last_name":"Sukhova","full_name":"Sukhova, Maria M."},{"first_name":"Artem A.","last_name":"Mikhailov","full_name":"Mikhailov, Artem A."},{"full_name":"Prilipov, Alexey G.","last_name":"Prilipov","first_name":"Alexey G."},{"full_name":"Gorchakov, Andrey A.","last_name":"Gorchakov","first_name":"Andrey A."},{"first_name":"Alexander V.","full_name":"Filatov, Alexander V.","last_name":"Filatov"}],"publication":"npj Vaccines","scopus_import":"1","quality_controlled":"1","date_created":"2023-01-12T12:02:54Z","article_processing_charge":"No","month":"11","department":[{"_id":"FyKo"}],"language":[{"iso":"eng"}],"volume":7,"publication_status":"published"},{"article_processing_charge":"No","date_created":"2023-01-16T09:56:57Z","quality_controlled":"1","publication_status":"published","department":[{"_id":"LeSa"}],"volume":13,"language":[{"iso":"eng"}],"month":"09","title":"Isolation of an escape-resistant SARS-CoV-2 neutralizing nanobody from a novel synthetic nanobody library","oa":1,"publication_identifier":{"issn":["1664-3224"]},"pmid":1,"type":"journal_article","doi":"10.3389/fimmu.2022.965446","article_number":"965446","scopus_import":"1","publication":"Frontiers in Immunology","author":[{"last_name":"Dormeshkin","full_name":"Dormeshkin, Dmitri","first_name":"Dmitri"},{"first_name":"Michail","full_name":"Shapira, Michail","last_name":"Shapira"},{"first_name":"Simon","last_name":"Dubovik","full_name":"Dubovik, Simon"},{"full_name":"Kavaleuski, Anton","last_name":"Kavaleuski","id":"4968f7ad-eb97-11eb-a6c2-8ed382e8912c","first_name":"Anton","orcid":"0000-0003-2091-526X"},{"full_name":"Katsin, Mikalai","last_name":"Katsin","first_name":"Mikalai"},{"full_name":"Migas, Alexandr","last_name":"Migas","first_name":"Alexandr"},{"first_name":"Alexander","full_name":"Meleshko, Alexander","last_name":"Meleshko"},{"first_name":"Sergei","last_name":"Semyonov","full_name":"Semyonov, Sergei"}],"external_id":{"isi":["000862479100001"],"pmid":["36189235"]},"intvolume":"        13","file":[{"file_id":"12443","success":1,"date_created":"2023-01-30T09:22:26Z","file_size":5695892,"file_name":"2022_FrontiersImmunology_Dormeshkin.pdf","relation":"main_file","access_level":"open_access","checksum":"f8f5d8110710033d0532e7e08bf9dad4","creator":"dernst","date_updated":"2023-01-30T09:22:26Z","content_type":"application/pdf"}],"status":"public","_id":"12252","oa_version":"Published Version","isi":1,"day":"16","publisher":"Frontiers Media","year":"2022","acknowledgement":"The authors declare that this study received funding from Immunofusion. The funder was not involved in the study design, collection, analysis, interpretation of data, the writing of this article or the decision to submit it for publication.","citation":{"ista":"Dormeshkin D, Shapira M, Dubovik S, Kavaleuski A, Katsin M, Migas A, Meleshko A, Semyonov S. 2022. Isolation of an escape-resistant SARS-CoV-2 neutralizing nanobody from a novel synthetic nanobody library. Frontiers in Immunology. 13, 965446.","ama":"Dormeshkin D, Shapira M, Dubovik S, et al. Isolation of an escape-resistant SARS-CoV-2 neutralizing nanobody from a novel synthetic nanobody library. <i>Frontiers in Immunology</i>. 2022;13. doi:<a href=\"https://doi.org/10.3389/fimmu.2022.965446\">10.3389/fimmu.2022.965446</a>","ieee":"D. Dormeshkin <i>et al.</i>, “Isolation of an escape-resistant SARS-CoV-2 neutralizing nanobody from a novel synthetic nanobody library,” <i>Frontiers in Immunology</i>, vol. 13. Frontiers Media, 2022.","apa":"Dormeshkin, D., Shapira, M., Dubovik, S., Kavaleuski, A., Katsin, M., Migas, A., … Semyonov, S. (2022). Isolation of an escape-resistant SARS-CoV-2 neutralizing nanobody from a novel synthetic nanobody library. <i>Frontiers in Immunology</i>. Frontiers Media. <a href=\"https://doi.org/10.3389/fimmu.2022.965446\">https://doi.org/10.3389/fimmu.2022.965446</a>","chicago":"Dormeshkin, Dmitri, Michail Shapira, Simon Dubovik, Anton Kavaleuski, Mikalai Katsin, Alexandr Migas, Alexander Meleshko, and Sergei Semyonov. “Isolation of an Escape-Resistant SARS-CoV-2 Neutralizing Nanobody from a Novel Synthetic Nanobody Library.” <i>Frontiers in Immunology</i>. Frontiers Media, 2022. <a href=\"https://doi.org/10.3389/fimmu.2022.965446\">https://doi.org/10.3389/fimmu.2022.965446</a>.","short":"D. Dormeshkin, M. Shapira, S. Dubovik, A. Kavaleuski, M. Katsin, A. Migas, A. Meleshko, S. Semyonov, Frontiers in Immunology 13 (2022).","mla":"Dormeshkin, Dmitri, et al. “Isolation of an Escape-Resistant SARS-CoV-2 Neutralizing Nanobody from a Novel Synthetic Nanobody Library.” <i>Frontiers in Immunology</i>, vol. 13, 965446, Frontiers Media, 2022, doi:<a href=\"https://doi.org/10.3389/fimmu.2022.965446\">10.3389/fimmu.2022.965446</a>."},"date_published":"2022-09-16T00:00:00Z","has_accepted_license":"1","ddc":["570"],"tmp":{"image":"/images/cc_by.png","short":"CC BY (4.0)","legal_code_url":"https://creativecommons.org/licenses/by/4.0/legalcode","name":"Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)"},"keyword":["Immunology","Immunology and Allergy","COVID-19","SARS-CoV-2","synthetic library","RBD","neutralization nanobody","VHH"],"fulldoi":"https://doi.org/10.3389/fimmu.2022.965446","article_type":"original","date_updated":"2025-06-11T13:42:26Z","user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","file_date_updated":"2023-01-30T09:22:26Z","abstract":[{"text":"The COVID−19 pandemic not only resulted in a global crisis, but also accelerated vaccine development and antibody discovery. Herein we report a synthetic humanized VHH library development pipeline for nanomolar-range affinity VHH binders to SARS-CoV-2 variants of concern (VoC) receptor binding domains (RBD) isolation. Trinucleotide-based randomization of CDRs by Kunkel mutagenesis with the subsequent rolling-cycle amplification resulted in more than 10<jats:sup>11</jats:sup> diverse phage display library in a manageable for a single person number of electroporation reactions. We identified a number of nanomolar-range affinity VHH binders to SARS-CoV-2 variants of concern (VoC) receptor binding domains (RBD) by screening a novel synthetic humanized antibody library. In order to explore the most robust and fast method for affinity improvement, we performed affinity maturation by CDR1 and CDR2 shuffling and avidity engineering by multivalent trimeric VHH fusion protein construction. As a result, H7-Fc and G12x3-Fc binders were developed with the affinities in nM and pM range respectively. Importantly, these affinities are weakly influenced by most of SARS-CoV-2 VoC mutations and they retain moderate binding to BA.4\\5. The plaque reduction neutralization test (PRNT) resulted in IC50 = 100 ng\\ml and 9.6 ng\\ml for H7-Fc and G12x3-Fc antibodies, respectively, for the emerging Omicron BA.1 variant. Therefore, these VHH could expand the present landscape of SARS-CoV-2 neutralization binders with the therapeutic potential for present and future SARS-CoV-2 variants.","lang":"eng"}]}]