{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,2]],"date-time":"2026-10-02T20:30:04Z","timestamp":1790973004719,"version":"4.1.0"},"reference-count":48,"publisher":"Oxford University Press (OUP)","issue":"4","license":[{"start":{"date-parts":[[2022,9,4]],"date-time":"2022-09-04T00:00:00Z","timestamp":1662249600000},"content-version":"vor","delay-in-days":3,"URL":"https:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"publisher","award":["S10OD021600"],"award-info":[{"award-number":["S10OD021600"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"publisher","award":["P41GM103832"],"award-info":[{"award-number":["P41GM103832"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"publisher","award":["P01GM121203"],"award-info":[{"award-number":["P01GM121203"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"publisher","award":["DP2HL137186"],"award-info":[{"award-number":["DP2HL137186"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2022,9,1]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>Host cell invasion by intracellular, eukaryotic parasites within the phylum Apicomplexa is a remarkable and active process involving the coordinated action of apical organelles and other structures. To date, capturing how these structures interact during invasion has been difficult to observe in detail. Here, we used cryogenic electron tomography to image the apical complex of Toxoplasma gondii tachyzoites under conditions that mimic resting parasites and those primed to invade through stimulation with calcium ionophore. Through the application of mixed-scale dense networks for image processing, we developed a highly efficient pipeline for annotation of tomograms, enabling us to identify and extract densities of relevant subcellular organelles and accurately analyze features in 3-D. The results reveal a dramatic change in the shape of the anteriorly located apical vesicle upon its apparent fusion with a rhoptry that occurs only in the stimulated parasites. We also present information indicating that this vesicle originates from the vesicles that parallel the intraconoidal microtubules and that the latter two structures are linked by a novel tether. We show that a rosette structure previously proposed to be involved in rhoptry secretion is associated with apical vesicles beyond just the most anterior one. This result, suggesting multiple vesicles are primed to enable rhoptry secretion, may shed light on the mechanisms Toxoplasma employs to enable repeated invasion attempts. Using the same approach, we examine Plasmodium falciparum merozoites and show that they too possess an apical vesicle just beneath a rosette, demonstrating evolutionary conservation of this overall subcellular organization.<\/jats:p>","DOI":"10.1093\/pnasnexus\/pgac183","type":"journal-article","created":{"date-parts":[[2022,9,4]],"date-time":"2022-09-04T04:59:07Z","timestamp":1662267547000},"source":"Crossref","is-referenced-by-count":34,"title":["Cryo-electron tomography with mixed-scale dense neural networks reveals key steps in deployment of\n                    <i>Toxoplasma<\/i>\n                    invasion machinery"],"prefix":"10.1093","volume":"1","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-4151-5513","authenticated-orcid":false,"given":"Li-av","family":"Segev-Zarko","sequence":"first","affiliation":[{"name":"Department of Microbiology and Immunology, Stanford University School of Medicine , 291 Campus Drive, Stanford, CA 94305 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4452-2663","authenticated-orcid":false,"given":"Peter D","family":"Dahlberg","sequence":"additional","affiliation":[{"name":"Department of Chemistry, Stanford University , 450 Serra Mall, Stanford, CA 94305 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Stella Y","family":"Sun","sequence":"additional","affiliation":[{"name":"Department of Bioengineering, Stanford University , 450 Serra Mall, Stanford, CA 94305 , USA"},{"name":"Department of Structural Biology, University of Pittsburgh , 4200 Fifth Ave, Pittsburgh, PA 15260 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dani\u00ebl M","family":"Pelt","sequence":"additional","affiliation":[{"name":"Leiden Institute of Advanced Computer Science, Leiden University , Rapenburg 70, 2311 EZ Leiden , The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chi Yong","family":"Kim","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Immunology, Stanford University School of Medicine , 291 Campus Drive, Stanford, CA 94305 , USA"},{"name":"Department of Pediatrics\u2013\u2013Infectious Diseases, Stanford University School of Medicine , 291 Campus Drive, Stanford, CA 94305 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2112-7700","authenticated-orcid":false,"given":"Elizabeth S","family":"Egan","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Immunology, Stanford University School of Medicine , 291 Campus Drive, Stanford, CA 94305 , USA"},{"name":"Department of Pediatrics\u2013\u2013Infectious Diseases, Stanford University School of Medicine , 291 Campus Drive, Stanford, CA 94305 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"James A","family":"Sethian","sequence":"additional","affiliation":[{"name":"Department of Mathematics, University of California , Berkeley, CA 94720 , USA"},{"name":"Center for Advanced Mathematics for Energy Research Application (CAMERA), Lawrence Berkeley National Laboratory , 1 Cyclotron Rd, Berkeley, CA 94720 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8910-3078","authenticated-orcid":false,"given":"Wah","family":"Chiu","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Immunology, Stanford University School of Medicine , 291 Campus Drive, Stanford, CA 94305 , USA"},{"name":"Department of Bioengineering, Stanford University , 450 Serra Mall, Stanford, CA 94305 , USA"},{"name":"Division of Cryo-EM and Bioimaging, SSRL, SLAC National Accelerator Laboratory, Stanford University , Menlo Park, CA 94305 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"John C","family":"Boothroyd","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Immunology, Stanford University School of Medicine , 291 Campus Drive, Stanford, CA 94305 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"286","published-online":{"date-parts":[[2022,9,4]]},"reference":[{"issue":"10195","key":"2023012507052534500_bib1","doi-asserted-by":"crossref","first-page":"332","DOI":"10.1016\/S0140-6736(19)31096-7","article-title":"Mapping the global endemicity and clinical burden of Plasmodium vivax, 2000\u201317: a spatial and temporal modelling study","volume":"394","author":"Battle","year":"2019","journal-title":"Lancet"},{"issue":"3","key":"2023012507052534500_bib2","doi-asserted-by":"crossref","first-page":"281","DOI":"10.4103\/0974-777X.83536","article-title":"Toxoplasmosis: a global threat","volume":"3","author":"Furtado","year":"2011","journal-title":"J Glob Infect Dis"},{"issue":"1","key":"2023012507052534500_bib3","first-page":"1","article-title":"Global, regional, and country seroprevalence of Toxoplasma gondiiin pregnant women: a systematic review, modelling and meta-analysis","volume":"10","author":"Bigna","year":"2020","journal-title":"Sci Rep 2020"},{"issue":"1","key":"2023012507052534500_bib4","doi-asserted-by":"crossref","first-page":"21","DOI":"10.1128\/MMBR.66.1.21-38.2002","article-title":"Cytoskeleton of apicomplexan parasites","volume":"66","author":"Morrissette","year":"2002","journal-title":"Microbiol Mol Biol Rev"},{"key":"2023012507052534500_bib5","doi-asserted-by":"crossref","first-page":"688","DOI":"10.1016\/j.pt.2020.05.001","article-title":"Evolution, composition, assembly, and function of the conoid in Apicomplexa","volume":"36","author":"Dos\u00a0Santos\u00a0Pacheco","year":"2020","journal-title":"Trends Parasitol"},{"issue":"1","key":"2023012507052534500_bib6","doi-asserted-by":"crossref","first-page":"83","DOI":"10.1016\/j.mib.2006.06.017","article-title":"Pulling together: an integrated model of Toxoplasma cell invasion","volume":"10","author":"Carruthers","year":"2007","journal-title":"Curr Opin Microbiol"},{"issue":"6","key":"2023012507052534500_bib7","doi-asserted-by":"crossref","first-page":"1039","DOI":"10.1083\/jcb.200112086","article-title":"A novel polymer of tubulin forms the conoid of Toxoplasma gondii","volume":"156","author":"Hu","year":"2002","journal-title":"J Cell Biol"},{"issue":"6","key":"2023012507052534500_bib8","doi-asserted-by":"crossref","first-page":"e2111661119","DOI":"10.1073\/pnas.2111661119","article-title":"Cryo-ET of Toxoplasmaparasites gives subnanometer insight into tubulin-based structures","volume":"119","author":"Sun","year":"2022","journal-title":"Proc Natl Acad Sci"},{"issue":"2","key":"2023012507052534500_bib9","doi-asserted-by":"crossref","first-page":"e13","DOI":"10.1371\/journal.ppat.0020013","article-title":"Cytoskeletal components of an invasion machine\u2014the apical complex of Toxoplasma gondii","volume":"2","author":"Hu","year":"2006","journal-title":"PLoS Pathog"},{"issue":"2","key":"2023012507052534500_bib10","doi-asserted-by":"crossref","first-page":"217","DOI":"10.1111\/j.1550-7408.1987.tb03162.x","article-title":"Cytoskeleton of Toxoplasma gondii","volume":"34","author":"Nichols","year":"1987","journal-title":"J Protozool"},{"issue":"5","key":"2023012507052534500_bib11","doi-asserted-by":"crossref","first-page":"e13018","DOI":"10.1111\/cmi.13018","article-title":"Biogenesis and secretion of micronemes in Toxoplasma gondii","volume":"21","author":"Dubois","year":"2019","journal-title":"Cell Microbiol"},{"issue":"3","key":"2023012507052534500_bib12","doi-asserted-by":"crossref","first-page":"453","DOI":"10.1111\/mmi.14674","article-title":"Biogenesis and discharge of the rhoptries: key organelles for entry and hijack of host cells by the Apicomplexa","volume":"115","author":"Ben\u00a0Chaabene","year":"2021","journal-title":"Mol Microbiol"},{"issue":"2","key":"2023012507052534500_bib13","doi-asserted-by":"crossref","first-page":"e17","DOI":"10.1371\/journal.ppat.0010017","article-title":"Identification of the moving junction complex of Toxoplasma gondii: a collaboration between distinct secretory organelles","volume":"1","author":"Alexander","year":"2005","journal-title":"PLOS Pathog"},{"key":"2023012507052534500_bib14","doi-asserted-by":"crossref","first-page":"420","DOI":"10.1016\/j.jsb.2011.11.028","article-title":"Dynamics and 3D organization of secretory organelles of Toxoplasma gondii","volume":"177","author":"Paredes-Santos","year":"2012","journal-title":"J Struct Biol"},{"issue":"4","key":"2023012507052534500_bib15","first-page":"425","article-title":"An Alveolata secretory machinery adapted to parasite host cell invasion","volume":"6","author":"Aquilini","year":"2021","journal-title":"Nat Microbiol 2021 64"},{"issue":"3","key":"2023012507052534500_bib16","doi-asserted-by":"crossref","first-page":"583","DOI":"10.2307\/3276331","article-title":"Electron microscope study of the proliferative form of Besnoitiajellisoni","volume":"52","author":"Sheffield","year":"1966","journal-title":"J Parasitol"},{"issue":"2","key":"2023012507052534500_bib17","doi-asserted-by":"crossref","first-page":"94","DOI":"10.1016\/S0022-5320(78)90012-6","article-title":"Freeze fracture study of the pellicle of an Eimerian sporozoite (Protozoa, Coccidia)","volume":"62","author":"Dubremetz","year":"1978","journal-title":"J Ultrastruct Res"},{"issue":"2","key":"2023012507052534500_bib18","doi-asserted-by":"crossref","first-page":"194","DOI":"10.1016\/S0022-5320(83)90130-2","article-title":"Are rhoptries of Coccidiareally extrusomes?","volume":"84","author":"Porchet-Hennere","year":"1983","journal-title":"J Ultrastruct Res"},{"issue":"1","key":"2023012507052534500_bib19","first-page":"1","article-title":"In situ ultrastructures of two evolutionarily distant apicomplexan rhoptry secretion systems","volume":"12","author":"Mageswaran","year":"2021","journal-title":"Nat Commun"},{"issue":"1","key":"2023012507052534500_bib20","doi-asserted-by":"crossref","first-page":"57","DOI":"10.3390\/cells8010057","article-title":"Cellular and structural studies of eukaryotic cells by cryo-electron tomography","volume":"8","author":"Weber","year":"2019","journal-title":"Cells"},{"issue":"2","key":"2023012507052534500_bib21","doi-asserted-by":"crossref","first-page":"120","DOI":"10.1111\/j.1550-7408.1996.tb04491.x","article-title":"Ca(2+)-dependence of conoid extrusion in Toxoplasma gondii tachyzoites","volume":"43","author":"Mondragon","year":"1996","journal-title":"J Eukaryot Microbiol"},{"issue":"2","key":"2023012507052534500_bib22","doi-asserted-by":"crossref","first-page":"421","DOI":"10.1046\/j.1365-2958.1999.01174.x","article-title":"Mobilization of intracellular calcium stimulates microneme discharge in Toxoplasma gondii","volume":"31","author":"Carruthers","year":"1999","journal-title":"Mol Microbiol"},{"issue":"24","key":"2023012507052534500_bib23","doi-asserted-by":"crossref","first-page":"9399","DOI":"10.1128\/MCB.20.24.9399-9408.2000","article-title":"Ionophore-resistant mutants of Toxoplasma gondiireveal host cell permeabilization as an early event in egress","volume":"20","author":"Black","year":"2000","journal-title":"Mol Cell Biol"},{"issue":"1","key":"2023012507052534500_bib24","first-page":"92","article-title":"Involvement of calcium and calmodulin in Toxoplasma gondiitachyzoite invasion","volume":"74","author":"Pezzella","year":"1997","journal-title":"Eur J Cell Biol"},{"issue":"2","key":"2023012507052534500_bib25","doi-asserted-by":"crossref","first-page":"254","DOI":"10.1073\/pnas.1715832114","article-title":"A mixed-scale dense convolutional neural network for image analysis","volume":"115","author":"Pelt","year":"2017","journal-title":"Proc Natl Acad Sci USA"},{"issue":"3","key":"2023012507052534500_bib26","doi-asserted-by":"crossref","first-page":"291","DOI":"10.1111\/j.1550-7408.1987.tb03177.x","article-title":"Effects of extracellular potassium on acid release and motility initiation in Toxoplasma gondii","volume":"34","author":"Endo","year":"1987","journal-title":"J Protozool"},{"issue":"44","key":"2023012507052534500_bib27","doi-asserted-by":"crossref","first-page":"41492","DOI":"10.1074\/jbc.M106154200","article-title":"The Loss of Cytoplasmic Potassium upon Host Cell Breakdown Triggers Egress of Toxoplasma gondii","volume":"276","author":"Moudy","year":"2001","journal-title":"J Biol Chem"},{"key":"2023012507052534500_bib28","volume-title":"U-net: Convolutional Networks for Biomedical Image Segmentation","author":"Ronneberger","year":"2015"},{"issue":"12","key":"2023012507052534500_bib29","doi-asserted-by":"crossref","first-page":"2481","DOI":"10.1109\/TPAMI.2016.2644615","article-title":"SegNet: a deep convolutional encoder-decoder architecture for image segmentation","volume":"39","author":"Badrinarayanan","year":"2017","journal-title":"IEEE Trans Pattern Anal Mach Intell"},{"key":"2023012507052534500_bib30","first-page":"063628","article-title":"Toxoplasma ferlin1 is a versatile and dynamic mediator of microneme trafficking and secretion","author":"Tagoe","year":"2020","journal-title":"bioRxiv"},{"issue":"6065","key":"2023012507052534500_bib31","doi-asserted-by":"crossref","first-page":"218","DOI":"10.1126\/science.1210829","article-title":"A DOC2 protein identified by mutational profiling is essential for apicomplexan parasite exocytosis","volume":"335","author":"Farrell","year":"2012","journal-title":"Science"},{"issue":"1","key":"2023012507052534500_bib32","doi-asserted-by":"crossref","first-page":"42","DOI":"10.1007\/s12035-009-8094-8","article-title":"DOC2B, C2 domains, and calcium: a tale of intricate interactions","volume":"41","author":"Friedrich","year":"2010","journal-title":"Mol Neurobiol"},{"issue":"6","key":"2023012507052534500_bib33","doi-asserted-by":"crossref","first-page":"1295","DOI":"10.1083\/jcb.113.6.1295","article-title":"Quenched flow analysis of exocytosis in Paramecium cells: time course, changes in membrane structure, and calcium requirements revealed after rapid mixing and rapid freezing of intact cells","volume":"113","author":"Knoll","year":"1991","journal-title":"J Cell Biol"},{"issue":"3","key":"2023012507052534500_bib34","doi-asserted-by":"crossref","first-page":"181","DOI":"10.1006\/jsbi.2002.4444","article-title":"Morphological changes during conoid extrusion in Toxoplasma gondiitachyzoites treated with calcium ionophore","volume":"136","author":"Monteiro","year":"2001","journal-title":"J Struct Biol"},{"key":"2023012507052534500_bib35","first-page":"448283","article-title":"Cellular electron tomography of the apical complex in the apicomplexan parasite Eimeria tenellashows a highly organised gateway for regulated secretion","author":"Burrell","year":"2021","journal-title":"bioRxiv"},{"key":"2023012507052534500_bib36","doi-asserted-by":"crossref","first-page":"1230","DOI":"10.1038\/s41564-022-01171-3","article-title":"Rhoptry secretion system structure and priming in Plasmodium falciparumrevealed using in situ cryo-electron tomography","volume":"7","author":"Martinez","year":"2022","journal-title":"Nat Microbiol"},{"issue":"5","key":"2023012507052534500_bib37","doi-asserted-by":"crossref","first-page":"e01510","DOI":"10.1128\/mBio.01510-18","article-title":"A member of the Ferlin Calcium Sensor Family is essential for Toxoplasma gondiiRhoptry Secretion","volume":"9","author":"Coleman","year":"2018","journal-title":"MBio"},{"issue":"7","key":"2023012507052534500_bib38","doi-asserted-by":"crossref","first-page":"e1002825","DOI":"10.1371\/journal.ppat.1002825","article-title":"Toxoplasma co-opts host cells it does not invade","volume":"8","author":"Koshy","year":"2012","journal-title":"PLOS Pathog"},{"issue":"4","key":"2023012507052534500_bib39","doi-asserted-by":"crossref","first-page":"841","DOI":"10.1111\/j.1462-5822.2007.00909.x","article-title":"Rhoptries are major players in Toxoplasma gondiiinvasion and host cell interaction","volume":"9","author":"Dubremetz","year":"2007","journal-title":"Cell Microbiol"},{"issue":"11","key":"2023012507052534500_bib40","doi-asserted-by":"crossref","first-page":"1231","DOI":"10.1016\/j.str.2020.07.017","article-title":"Multi-scale 3D cryo-correlative microscopy for vitrified cells","volume":"28","author":"Wu","year":"2020","journal-title":"Structure"},{"key":"2023012507052534500_bib41","doi-asserted-by":"crossref","first-page":"253","DOI":"10.1146\/annurev-physchem-090319-051546","article-title":"Cryogenic super-resolution fluorescence and electron microscopy correlated at the nanoscale","volume":"72","author":"Dahlberg","year":"2021","journal-title":"Annu Rev Phys Chem"},{"issue":"32","key":"2023012507052534500_bib42","doi-asserted-by":"crossref","first-page":"14378","DOI":"10.1073\/pnas.1009198107","article-title":"Isolation of viable Plasmodium falciparummerozoites to define erythrocyte invasion events and advance vaccine and drug development","volume":"107","author":"Boyle","year":"2010","journal-title":"Proc Natl Acad Sci USA"},{"issue":"1","key":"2023012507052534500_bib43","doi-asserted-by":"crossref","first-page":"95","DOI":"10.1111\/j.1462-5822.2008.01242.x","article-title":"Irreversible effect of cysteine protease inhibitors on the release of malaria parasites from infected erythrocytes","volume":"11","author":"Glushakova","year":"2009","journal-title":"Cell Microbiol"},{"issue":"44","key":"2023012507052534500_bib44","doi-asserted-by":"crossref","first-page":"15635","DOI":"10.1073\/pnas.1418377111","article-title":"Volta potential phase plate for in-focus phase contrast transmission electron microscopy","volume":"111","author":"Danev","year":"2014","journal-title":"Proc Natl Acad Sci USA"},{"issue":"4","key":"2023012507052534500_bib45","doi-asserted-by":"crossref","first-page":"331","DOI":"10.1038\/nmeth.4193","article-title":"MotionCor2: anisotropic correction of beam-induced motion for improved cryo-electron microscopy","volume":"14","author":"Zheng","year":"2017","journal-title":"Nat Methods"},{"issue":"11","key":"2023012507052534500_bib46","doi-asserted-by":"crossref","first-page":"1161","DOI":"10.1038\/s41592-019-0591-8","article-title":"A complete data processing workflow for cryo-ET and subtomogram averaging","volume":"16","author":"Chen","year":"2019","journal-title":"Nat Methods"},{"issue":"3","key":"2023012507052534500_bib47","doi-asserted-by":"crossref","first-page":"343","DOI":"10.1006\/jsbi.1997.3919","article-title":"Dual-axis tomography: an approach with alignment methods that preserve resolution","volume":"120","author":"Mastronarde","year":"1997","journal-title":"J Struct Biol"},{"issue":"1","key":"2023012507052534500_bib48","doi-asserted-by":"crossref","first-page":"71","DOI":"10.1006\/jsbi.1996.0013","article-title":"Computer visualization of three-dimensional image data using IMOD","volume":"116","author":"Kremer","year":"1996","journal-title":"J Struct Biol"}],"container-title":["PNAS Nexus"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/academic.oup.com\/pnasnexus\/advance-article-pdf\/doi\/10.1093\/pnasnexus\/pgac183\/45688437\/pgac183.pdf","content-type":"application\/pdf","content-version":"am","intended-application":"syndication"},{"URL":"https:\/\/academic.oup.com\/pnasnexus\/article-pdf\/1\/4\/pgac183\/48849274\/pgac183.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/academic.oup.com\/pnasnexus\/article-pdf\/1\/4\/pgac183\/48849274\/pgac183.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,1,25]],"date-time":"2023-01-25T02:07:55Z","timestamp":1674612475000},"score":1,"resource":{"primary":{"URL":"https:\/\/academic.oup.com\/pnasnexus\/article\/doi\/10.1093\/pnasnexus\/pgac183\/6691727"}},"subtitle":[],"editor":[{"given":"Karen E","family":"Nelson","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"editor"}]}],"short-title":[],"issued":{"date-parts":[[2022,9,1]]},"references-count":48,"journal-issue":{"issue":"4","published-print":{"date-parts":[[2022,9,1]]}},"URL":"https:\/\/doi.org\/10.1093\/pnasnexus\/pgac183","relation":{"has-preprint":[{"id-type":"doi","id":"10.1101\/2022.01.12.476068","asserted-by":"object"}]},"ISSN":["2752-6542"],"issn-type":[{"value":"2752-6542","type":"electronic"}],"subject":[],"published-other":{"date-parts":[[2022,9]]},"published":{"date-parts":[[2022,9,1]]},"article-number":"pgac183"}}