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Here we solved these challenges by developing an integrated optical/machine learning technology, LIONESS (Live Information-Optimized Nanoscopy Enabling Saturated Segmentation). It leverages optical modifications to stimulated emission depletion (STED) microscopy in comprehensively, extracellularly labelled tissue and prior information on sample structure via machine learning to simultaneously achieve isotropic super-resolution, high signal-to-noise-ratio, and compatibility with living tissue. This allows dense deep-learning-based instance segmentation and 3D-reconstruction at synapse level incorporating molecular, activity, and morphodynamic information. LIONESS opens up avenues for studying the dynamic functional (nano-)architecture of living brain tissue."}],"license":"https://creativecommons.org/licenses/by-sa/4.0/","_id":"12817","acknowledgement":"We thank J. Vorlaufer, N. Agudelo, A. Wartak for microscope maintenance and troubleshooting, C. Kreuzinger and A. Freeman for technical assistance, and M. Šuplata for hardware control support, and Márcia Cunha dos Santos for initial exploration of software. We thank Paul Henderson for advice on deep-learning training and Michael Sixt, Scott Boyd, and Tamara Weiss for discussions and critical reading of the manuscript. Luke Lavis (Janelia Research Campus) generously provided JF585-HaloTag ligand. 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PSD95-HaloTag mice were kindly provided by Seth Grant (University of Edinburgh). We acknowledge expert support by IST Austria’s scientific computing, imaging and optics, preclinical, and lab support facilities, and by the Library and Miba machine shop.\r\nWe gratefully acknowledge funding by the following sources: \r\nAustrian Science Fund (FWF) grant I3600-B27 (JGD)\r\nAustrian Science Fund (FWF) grant DK W1232 (JGD, JMM)\r\nAustrian Science Fund (FWF) grant Z 312-B27, Wittgenstein award (PJ)\r\nAustrian Science Funds (FWF) projects I4685-B, I6565-B (SYNABS) and DOC 33-B27 (RH)\r\nGesellschaft für Forschungsförderung NÖ (NFB) grant LSC18-022 (JGD)\r\nEuropean Union’s Horizon 2020 research and innovation programme, European Research Council (ERC) grant 715508 – REVERSEAUTISM (GN)\r\nEuropean Union’s Horizon 2020 research and innovation programme, European Research Council (ERC) grant 692692 – GIANTSYN (PJ)\r\nMarie Skłodowska-Curie Actions Fellowship GA no. 665385 under the EU Horizon 2020 program (JMM, JL)\r\nMarie Skłodowska-Curie Actions Individual Fellowship 101026635 under the EU Horizon 2020 program (JFW)","year":"2023","department":[{"_id":"JoDa"},{"_id":"SaSi"},{"_id":"GaNo"},{"_id":"PeJo"},{"_id":"Bio"},{"_id":"RySh"}],"publisher":"Institute of Science and Technology Austria","author":[{"full_name":"Danzl, Johann G","last_name":"Danzl","first_name":"Johann G","orcid":"0000-0001-8559-3973","id":"42EFD3B6-F248-11E8-B48F-1D18A9856A87"}],"contributor":[{"last_name":"Michalska","first_name":"Julia M","id":"443DB6DE-F248-11E8-B48F-1D18A9856A87"},{"last_name":"Lyudchik","first_name":"Julia","id":"46E28B80-F248-11E8-B48F-1D18A9856A87"},{"last_name":"Velicky","first_name":"Philipp","orcid":"0000-0002-2340-7431","id":"39BDC62C-F248-11E8-B48F-1D18A9856A87"},{"last_name":"Stefanickova","first_name":"Hana","id":"ee3cb6ca-ec98-11ea-ae11-ff703e2254ed"},{"id":"63836096-4690-11EA-BD4E-32803DDC885E","orcid":"0000-0002-8698-3823","first_name":"Jake","last_name":"Watson"},{"id":"9ac8f577-2357-11eb-997a-e566c5550886","last_name":"Cenameri","first_name":"Alban"},{"last_name":"Sommer","first_name":"Christoph M","orcid":"0000-0003-1216-9105","id":"4DF26D8C-F248-11E8-B48F-1D18A9856A87"},{"first_name":"Nicole","last_name":"Amberg","id":"4CD6AAC6-F248-11E8-B48F-1D18A9856A87","orcid":"0000-0002-3183-8207"},{"orcid":"0000-0003-2356-9403","id":"41CB84B2-F248-11E8-B48F-1D18A9856A87","last_name":"Venturino","first_name":"Alessandro"},{"first_name":"Karl","last_name":"Roessler"},{"last_name":"Czech","first_name":"Thomas"},{"last_name":"Höftberger","first_name":"Romana"},{"last_name":"Siegert","first_name":"Sandra","orcid":"0000-0001-8635-0877","id":"36ACD32E-F248-11E8-B48F-1D18A9856A87"},{"id":"3E57A680-F248-11E8-B48F-1D18A9856A87","orcid":"0000-0002-7673-7178","first_name":"Gaia","last_name":"Novarino"},{"orcid":"0000-0001-5001-4804","id":"353C1B58-F248-11E8-B48F-1D18A9856A87","last_name":"Jonas","first_name":"Peter M"}],"related_material":{"record":[{"relation":"used_in_publication","status":"public","id":"14257"}],"link":[{"relation":"research_data","description":"Original data for Fig. 5d, Fig. 5d (N2V) and Fig. 5f-i, provided via an external link due to the large size (>10GB) of the datasets. 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Chiral and Nematic Phases of Flexible Active Filaments. Institute of Science and Technology Austria, 2023, doi:10.15479/AT:ISTA:13116.","short":"Z. Dunajova, B. Prats Mateu, P. Radler, K. Lim, D. Brandis, P. Velicky, J.G. Danzl, R.W. Wong, J. Elgeti, E.B. Hannezo, M. Loose, (2023).","chicago":"Dunajova, Zuzana, Batirtze Prats Mateu, Philipp Radler, Keesiang Lim, Dörte Brandis, Philipp Velicky, Johann G Danzl, et al. “Chiral and Nematic Phases of Flexible Active Filaments.” Institute of Science and Technology Austria, 2023. https://doi.org/10.15479/AT:ISTA:13116.","ama":"Dunajova Z, Prats Mateu B, Radler P, et al. Chiral and nematic phases of flexible active filaments. 2023. doi:10.15479/AT:ISTA:13116","ista":"Dunajova Z, Prats Mateu B, Radler P, Lim K, Brandis D, Velicky P, Danzl JG, Wong RW, Elgeti J, Hannezo EB, Loose M. 2023. Chiral and nematic phases of flexible active filaments, Institute of Science and Technology Austria, 10.15479/AT:ISTA:13116.","ieee":"Z. Dunajova et al., “Chiral and nematic phases of flexible active filaments.” Institute of Science and Technology Austria, 2023.","apa":"Dunajova, Z., Prats Mateu, B., Radler, P., Lim, K., Brandis, D., Velicky, P., … Loose, M. (2023). Chiral and nematic phases of flexible active filaments. Institute of Science and Technology Austria. https://doi.org/10.15479/AT:ISTA:13116"},"oa":1,"project":[{"name":"Self-Organization of the Bacterial Cell","call_identifier":"H2020","_id":"2595697A-B435-11E9-9278-68D0E5697425","grant_number":"679239"},{"name":"Understanding bacterial cell division by in vitro\r\nreconstitution","grant_number":"P34607","_id":"fc38323b-9c52-11eb-aca3-ff8afb4a011d"},{"name":"Motile active matter models of migrating cells and chiral filaments","_id":"34d75525-11ca-11ed-8bc3-89b6307fee9d","grant_number":"26360"}],"abstract":[{"lang":"eng","text":"The emergence of large-scale order in self-organized systems relies on local interactions between individual components. During bacterial cell division, FtsZ -- a prokaryotic homologue of the eukaryotic protein tubulin -- polymerizes into treadmilling filaments that further organize into a cytoskeletal ring. In vitro, FtsZ filaments can form dynamic chiral assemblies. However, how the active and passive properties of individual filaments relate to these large-scale self-organized structures remains poorly understood. Here, we connect single filament properties with the mesoscopic scale by combining minimal active matter simulations and biochemical reconstitution experiments. We show that density and flexibility of active chiral filaments define their global order. At intermediate densities, curved, flexible filaments organize into chiral rings and polar bands. An effectively nematic organization dominates for high densities and for straight, mutant filaments with increased rigidity. Our predicted phase diagram captures these features quantitatively, demonstrating how the flexibility, density and chirality of active filaments affect their collective behaviour. Our findings shed light on the fundamental properties of active chiral matter and explain how treadmilling FtsZ filaments organize during bacterial cell division. 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work was supported by the European Research Council through grant ERC 2015-StG-679239 and by the Austrian Science Fund (FWF) StandAlone P34607 to M.L., B. P.M. was also supported by the Kanazawa University WPI- NanoLSI Bio-SPM collaborative research program. Z.D. has received funding from Doctoral Programme of the Austrian Academy of Sciences (OeAW): Grant agreement 26360. We thank Jan Brugues (MPI CBG, Dresden, Germany), Andela Saric (ISTA, Klosterneuburg, Austria), Daniel Pearce (Uni Geneva, Switzerland) for valuable scientific input and comments on the manuscript. We are also thankful for the support by the Scientific Service Units (SSU) of IST Austria through resources provided by the Imaging and Optics Facility (IOF) and the Lab Support Facility (LSF). ","_id":"13116","year":"2023","user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","status":"public","ddc":["539"],"title":"Chiral and nematic phases of flexible active filaments","department":[{"_id":"MaLo"},{"_id":"EdHa"},{"_id":"JoDa"}],"publisher":"Institute of Science and Technology Austria"},{"ddc":["576"],"status":"public","title":"The scorpionfly (Panorpa cognata) genome highlights conserved and derived features of the peculiar dipteran X chromosome","department":[{"_id":"BeVi"}],"publisher":"Institute of Science and Technology Austria","_id":"14614","year":"2023","user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","date_created":"2023-11-27T16:39:19Z","date_updated":"2024-02-21T12:18:35Z","file":[{"file_id":"14625","relation":"main_file","date_created":"2023-11-28T13:15:26Z","date_updated":"2023-11-28T13:15:26Z","success":1,"checksum":"cd0f13322b5156819ecaebd2bc8e7d12","file_name":"panorpaX.zip","access_level":"open_access","creator":"clasne","file_size":404968272,"content_type":"application/zip"},{"date_updated":"2023-11-30T14:16:59Z","date_created":"2023-11-30T14:16:59Z","checksum":"9ff600416577687a737cb3c96dfcb26c","success":1,"relation":"main_file","file_id":"14634","file_size":2625,"content_type":"text/plain","creator":"clasne","file_name":"panorpa_readme.txt","access_level":"open_access"}],"oa_version":"Published Version","author":[{"first_name":"Clementine","last_name":"Lasne","id":"02225f57-50d2-11eb-9ed8-8c92b9a34237","orcid":"0000-0002-1197-8616","full_name":"Lasne, Clementine"},{"full_name":"Elkrewi, Marwan N","last_name":"Elkrewi","first_name":"Marwan N","orcid":"0000-0002-5328-7231","id":"0B46FACA-A8E1-11E9-9BD3-79D1E5697425"}],"related_material":{"record":[{"id":"14613","relation":"used_in_publication","status":"public"}]},"contributor":[{"orcid":"0000-0002-5328-7231","id":"0B46FACA-A8E1-11E9-9BD3-79D1E5697425","last_name":"Elkrewi","contributor_type":"researcher","first_name":"Marwan N"}],"type":"research_data","abstract":[{"text":"Many insects carry an ancient X chromosome—the Drosophila Muller element F—that likely predates their origin. Interestingly, the X has undergone turnover in multiple fly species (Diptera) after being conserved for more than 450 My. The long evolutionary distance between Diptera and other sequenced insect clades makes it difficult to infer what could have contributed to this sudden increase in rate of turnover. Here, we produce the first genome and transcriptome of scorpionflies (genus Panorpa), an insect belonging to a long overlooked sister-order to Diptera: Mecoptera. Combining our genome assembly with genomic short-read data, we obtain genome coverage and identify X-linked super-scaffolds. We further perform a gene homology analysis between the Panorpa X and a closely related Diptera species, and we assess the conservation of the Panorpa X-linked gene content with that of more distantly related insect species. We explored the structure of the Panorpa X by determining its repeat content, GC content, and nucleotide diversity. Finally, we used RNAseq data to detect the presence of dosage compensation in somatic tissues, as well as to explore gene expression tissue-specificity, and sex-bias in gene expression. We find high conservation of gene content between the mecopteran X and the dipteran Muller F element, as well as several shared biological features, such as the presence of dosage compensation and a low amount of genetic diversity, consistent with a low recombination rate. However, the 2 homologous X chromosomes differ strikingly in their size and number of genes they carry. Our results therefore support a common ancestry of the mecopteran and ancestral dipteran X chromosomes, and suggest that Muller element F shrank in size and gene content after the split of Diptera and Mecoptera, which may have contributed to its turnover in dipteran insects.","lang":"eng"}],"file_date_updated":"2023-11-30T14:16:59Z","tmp":{"name":"Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)","legal_code_url":"https://creativecommons.org/licenses/by/4.0/legalcode","short":"CC BY (4.0)","image":"/images/cc_by.png"},"oa":1,"citation":{"short":"C. Lasne, M.N. Elkrewi, (2023).","mla":"Lasne, Clementine, and Marwan N. Elkrewi. The Scorpionfly (Panorpa Cognata) Genome Highlights Conserved and Derived Features of the Peculiar Dipteran X Chromosome. Institute of Science and Technology Austria, 2023, doi:10.15479/AT:ISTA:14614.","chicago":"Lasne, Clementine, and Marwan N Elkrewi. “The Scorpionfly (Panorpa Cognata) Genome Highlights Conserved and Derived Features of the Peculiar Dipteran X Chromosome.” Institute of Science and Technology Austria, 2023. https://doi.org/10.15479/AT:ISTA:14614.","ama":"Lasne C, Elkrewi MN. The scorpionfly (Panorpa cognata) genome highlights conserved and derived features of the peculiar dipteran X chromosome. 2023. doi:10.15479/AT:ISTA:14614","ieee":"C. Lasne and M. N. Elkrewi, “The scorpionfly (Panorpa cognata) genome highlights conserved and derived features of the peculiar dipteran X chromosome.” Institute of Science and Technology Austria, 2023.","apa":"Lasne, C., & Elkrewi, M. N. (2023). The scorpionfly (Panorpa cognata) genome highlights conserved and derived features of the peculiar dipteran X chromosome. Institute of Science and Technology Austria. https://doi.org/10.15479/AT:ISTA:14614","ista":"Lasne C, Elkrewi MN. 2023. The scorpionfly (Panorpa cognata) genome highlights conserved and derived features of the peculiar dipteran X chromosome, Institute of Science and Technology Austria, 10.15479/AT:ISTA:14614."},"doi":"10.15479/AT:ISTA:14614","date_published":"2023-12-01T00:00:00Z","keyword":["Panorpa","scorpionfly","genome","transcriptome"],"month":"12","day":"01","has_accepted_license":"1","article_processing_charge":"No"},{"date_created":"2024-02-26T08:37:57Z","date_updated":"2024-02-26T08:45:55Z","oa_version":"Published Version","author":[{"full_name":"Curk, Samo","id":"031eff0d-d481-11ee-8508-cd12a7a86e5b","orcid":"0000-0001-6160-9766","first_name":"Samo","last_name":"Curk"}],"related_material":{"record":[{"status":"public","relation":"used_in_publication","id":"15001"}]},"ddc":["570"],"status":"public","title":"aggregation_data","publisher":"Figshare","department":[{"_id":"AnSa"}],"_id":"15027","year":"2023","user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","license":"https://creativecommons.org/publicdomain/zero/1.0/","abstract":[{"lang":"eng","text":"This data repository underpins the paper, published in PNAS (doi pending) and bioarxiv (doi: https://doi.org/10.1101/2023.07.05.547777)."}],"type":"research_data_reference","date_published":"2023-12-13T00:00:00Z","tmp":{"short":"CC0 (1.0)","image":"/images/cc_0.png","legal_code_url":"https://creativecommons.org/publicdomain/zero/1.0/legalcode","name":"Creative Commons Public Domain Dedication (CC0 1.0)"},"main_file_link":[{"open_access":"1","url":"https://figshare.com/s/85798bba4ebc68d822ed"}],"oa":1,"citation":{"short":"S. Curk, (2023).","mla":"Curk, Samo. Aggregation_data. Figshare, 2023.","chicago":"Curk, Samo. “Aggregation_data.” Figshare, 2023.","ama":"Curk S. aggregation_data. 2023.","apa":"Curk, S. (2023). aggregation_data. Figshare.","ieee":"S. Curk, “aggregation_data.” Figshare, 2023.","ista":"Curk S. 2023. aggregation_data, Figshare."},"day":"13","month":"12","article_processing_charge":"No","has_accepted_license":"1"}]