{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,11,5]],"date-time":"2026-11-05T03:37:47Z","timestamp":1793849867462,"version":"4.6.1"},"reference-count":85,"publisher":"Springer Science and Business Media LLC","issue":"1","license":[{"start":{"date-parts":[[2017,7,18]],"date-time":"2017-07-18T00:00:00Z","timestamp":1500336000000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"},{"start":{"date-parts":[[2017,7,18]],"date-time":"2017-07-18T00:00:00Z","timestamp":1500336000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Sci Rep"],"abstract":"<jats:title>Abstract<\/jats:title><jats:p>Chikungunya virus (CHIKV), a mosquito-borne alphavirus, causes febrile disease, muscle and joint pain, which can become chronic in some individuals. The non-structural protein 3 (nsP3) plays essential roles during infection, but a complete understanding of its function is lacking. Here we used a microscopy-based approach to image CHIKV nsP3 inside human cells. The SNAP system consists of a self-labelling enzyme tag, which catalyses the covalent linking of exogenously supplemented synthetic ligands. Genetic insertion of this tag resulted in viable replicons and specific labelling while preserving the effect of nsP3 on stress granule responses and co-localisation with GTPase Activating Protein (SH3 domain) Binding Proteins (G3BPs). With sub-diffraction, three-dimensional, optical imaging, we visualised nsP3-positive structures with variable density and morphology, including high-density rod-like structures, large spherical granules, and small, low-density structures. Next, we confirmed the utility of the SNAP-tag for studying protein turnover by pulse-chase labelling. We also revealed an association of nsP3 with cellular lipid droplets and examined the spatial relationships between nsP3 and the non-structural protein 1 (nsP1). Together, our study provides a sensitive, specific, and versatile system for fundamental research into the individual functions of a viral non-structural protein during infection with a medically important arthropod-borne virus (arbovirus).<\/jats:p>","DOI":"10.1038\/s41598-017-05820-0","type":"journal-article","created":{"date-parts":[[2017,7,12]],"date-time":"2017-07-12T09:09:28Z","timestamp":1499850568000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":29,"title":["SNAP-tagged Chikungunya Virus Replicons Improve Visualisation of Non-Structural Protein 3 by Fluorescence Microscopy"],"prefix":"10.1038","volume":"7","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-1644-6464","authenticated-orcid":false,"given":"Roland","family":"Remenyi","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Grace C.","family":"Roberts","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Carsten","family":"Zothner","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andres","family":"Merits","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9821-1003","authenticated-orcid":false,"given":"Mark","family":"Harris","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2017,7,18]]},"reference":[{"key":"5820_CR1","doi-asserted-by":"publisher","first-page":"1231","DOI":"10.1056\/NEJMra1406035","volume":"372","author":"SC Weaver","year":"2015","unstructured":"Weaver, S. C. & Lecuit, M. Chikungunya virus and the global spread of a mosquito-borne disease. The New England journal of medicine \n                           372, 1231\u20131239, doi:10.1056\/NEJMra1406035 (2015).","journal-title":"The New England journal of medicine"},{"key":"5820_CR2","doi-asserted-by":"publisher","first-page":"e2137","DOI":"10.1371\/journal.pntd.0002137","volume":"7","author":"C Schilte","year":"2013","unstructured":"Schilte, C. et al. Chikungunya virus-associated long-term arthralgia: a 36-month prospective longitudinal study. PLoS neglected tropical diseases \n                           7, e2137, doi:10.1371\/journal.pntd.0002137 (2013).","journal-title":"PLoS neglected tropical diseases"},{"key":"5820_CR3","first-page":"105","volume":"16","author":"B Meason","year":"2014","unstructured":"Meason, B. & Paterson, R. Chikungunya, climate change, and human rights. Health and human rights \n                           16, 105\u2013112 (2014).","journal-title":"Health and human rights"},{"key":"5820_CR4","doi-asserted-by":"publisher","first-page":"2976","DOI":"10.1016\/j.vaccine.2016.03.076","volume":"34","author":"C Smalley","year":"2016","unstructured":"Smalley, C., Erasmus, J. H., Chesson, C. B. & Beasley, D. W. Status of research and development of vaccines for chikungunya. Vaccine \n                           34, 2976\u20132981, doi:10.1016\/j.vaccine.2016.03.076 (2016).","journal-title":"Vaccine"},{"key":"5820_CR5","doi-asserted-by":"publisher","first-page":"59","DOI":"10.1016\/j.antiviral.2015.06.017","volume":"121","author":"R Abdelnabi","year":"2015","unstructured":"Abdelnabi, R., Neyts, J. & Delang, L. Towards antivirals against chikungunya virus. Antiviral research \n                           121, 59\u201368, doi:10.1016\/j.antiviral.2015.06.017 (2015).","journal-title":"Antiviral research"},{"key":"5820_CR6","doi-asserted-by":"crossref","first-page":"491","DOI":"10.1128\/mr.58.3.491-562.1994","volume":"58","author":"JH Strauss","year":"1994","unstructured":"Strauss, J. H. & Strauss, E. G. The alphaviruses: gene expression, replication, and evolution. Microbiological reviews \n                           58, 491\u2013562 (1994).","journal-title":"Microbiological reviews"},{"key":"5820_CR7","doi-asserted-by":"publisher","first-page":"108","DOI":"10.1186\/1743-422X-5-108","volume":"5","author":"AE Firth","year":"2008","unstructured":"Firth, A. E., Chung, B. Y., Fleeton, M. N. & Atkins, J. F. Discovery of frameshifting in Alphavirus 6K resolves a 20-year enigma. Virology journal \n                           5, 108, doi:10.1186\/1743-422X-5-108 (2008).","journal-title":"Virology journal"},{"key":"5820_CR8","doi-asserted-by":"publisher","first-page":"8511","DOI":"10.1128\/JVI.00449-13","volume":"87","author":"JE Snyder","year":"2013","unstructured":"Snyder, J. E. et al. Functional characterization of the alphavirus TF protein. Journal of virology \n                           87, 8511\u20138523, doi:10.1128\/JVI.00449-13 (2013).","journal-title":"Journal of virology"},{"key":"5820_CR9","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1621485114","author":"RL McPherson","year":"2017","unstructured":"McPherson, R. L. et al. ADP-ribosylhydrolase activity of Chikungunya virus macrodomain is critical for virus replication and virulence. Proceedings of the National Academy of Sciences of the United States of America. doi:10.1073\/pnas.1621485114 (2017).","journal-title":"Proceedings of the National Academy of Sciences of the United States of America"},{"key":"5820_CR10","doi-asserted-by":"publisher","first-page":"41746","DOI":"10.1038\/srep41746","volume":"7","author":"L Eckei","year":"2017","unstructured":"Eckei, L. et al. The conserved macrodomains of the non-structural proteins of Chikungunya virus and other pathogenic positive strand RNA viruses function as mono-ADP-ribosylhydrolases. Scientific reports \n                           7, 41746, doi:10.1038\/srep41746 (2017).","journal-title":"Scientific reports"},{"key":"5820_CR11","doi-asserted-by":"publisher","first-page":"305","DOI":"10.1016\/j.virol.2009.03.031","volume":"388","author":"E Park","year":"2009","unstructured":"Park, E. & Griffin, D. E. The nsP3 macro domain is important for Sindbis virus replication in neurons and neurovirulence in mice. 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Journal of virology \n                           84, 11679\u201311695, doi:10.1128\/JVI.01441-10 (2010).","journal-title":"Journal of virology"},{"key":"5820_CR18","doi-asserted-by":"publisher","unstructured":"Fros, J. J. & Pijlman, G. P. Alphavirus Infection: Host Cell Shut-Off and Inhibition of Antiviral Responses. Viruses \n                           8, 10.3390\/v8060166 (2016).","DOI":"10.3390\/v8060166"},{"key":"5820_CR19","doi-asserted-by":"publisher","first-page":"11420","DOI":"10.1128\/JVI.01579-15","volume":"89","author":"B Thaa","year":"2015","unstructured":"Thaa, B. et al. Differential Phosphatidylinositol-3-Kinase-Akt-mTOR Activation by Semliki Forest and Chikungunya Viruses Is Dependent on nsP3 and Connected to Replication Complex Internalization. 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New World and Old World Alphaviruses Have Evolved to Exploit Different Components of Stress Granules, FXR and G3BP Proteins, for Assembly of Viral Replication Complexes. PLoS pathogens \n                           12, e1005810, doi:10.1371\/journal.ppat.1005810 (2016).","journal-title":"PLoS pathogens"},{"key":"5820_CR28","doi-asserted-by":"publisher","first-page":"16534","DOI":"10.1073\/pnas.1210418109","volume":"109","author":"G Shin","year":"2012","unstructured":"Shin, G. et al. Structural and functional insights into alphavirus polyprotein processing and pathogenesis. 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