The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infection

Autores
Chapman, Sean; Faulkner, Christine; Kaiserli, Eirini; Garcia-Mata, Carlos; Savenkov, Eugene I.; Roberts, Alison G.; Oparka, Karl J.; Christie, John M.
Año de publicación
2008
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Fluorescent proteins (FPs) based on green fluorescent protein (GFP) are widely used throughout cell biology to study protein dynamics, and have extensive use as reporters of virus infection and spread. However, FP-tagging of viruses is limited by the constraints of viral genome size resulting in FP loss through recombination events. To overcome this, we have engineered a smaller (10 kDa) flavinbased alternative to GFP (25 kDa) derived from the light, oxygen or voltage-sensing (LOV) domain of the plant blue light receptor, phototropin. Molecular evolution and Tobacco mosaic virus (TMV)- based expression screening produced LOV variants with improved fluorescence and photostability in planta. One variant in particular, designated iLOV, possessed photophysical properties that made it ideally suited as a reporter of subcellular protein localization in both plant and mammalian cells. Moreover, iLOV fluorescence was found to recover spontaneously after photobleaching and displayed an intrinsic photochemistry conferring advantages over GFP-based FPs. When expressed either as a cytosolic protein or as a viral protein fusion, iLOV functioned as a superior reporter to GFP for monitoring local and systemic infections of plant RNA viruses. iLOV, therefore, offers greater utility in FP-tagging of viral gene products and represents a viable alternative where functional protein expression is limited by steric constraints or genome size.
Fil: Chapman, Sean. Scottish Crop Research Institute; Reino Unido
Fil: Faulkner, Christine. University of Edinburgh; Reino Unido
Fil: Kaiserli, Eirini. University of Glasgow; Reino Unido
Fil: Garcia-Mata, Carlos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina. University of Glasgow; Reino Unido
Fil: Savenkov, Eugene I.. University of Agricultural Sciences; Suecia
Fil: Roberts, Alison G.. Scottish Crop Research Institute; Reino Unido
Fil: Oparka, Karl J.. University of Edinburgh; Reino Unido
Fil: Christie, John M.. University of Glasgow; Reino Unido
Materia
FLUORESCENCE IMAGING
LOV DOMAIN
MOLECULAR EVOLUTION
PHOTORECEPTOR
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/138316
Acceder
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network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infectionChapman, SeanFaulkner, ChristineKaiserli, EiriniGarcia-Mata, CarlosSavenkov, Eugene I.Roberts, Alison G.Oparka, Karl J.Christie, John M.FLUORESCENCE IMAGINGLOV DOMAINMOLECULAR EVOLUTIONPHOTORECEPTORhttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Fluorescent proteins (FPs) based on green fluorescent protein (GFP) are widely used throughout cell biology to study protein dynamics, and have extensive use as reporters of virus infection and spread. However, FP-tagging of viruses is limited by the constraints of viral genome size resulting in FP loss through recombination events. To overcome this, we have engineered a smaller (10 kDa) flavinbased alternative to GFP (25 kDa) derived from the light, oxygen or voltage-sensing (LOV) domain of the plant blue light receptor, phototropin. Molecular evolution and Tobacco mosaic virus (TMV)- based expression screening produced LOV variants with improved fluorescence and photostability in planta. One variant in particular, designated iLOV, possessed photophysical properties that made it ideally suited as a reporter of subcellular protein localization in both plant and mammalian cells. Moreover, iLOV fluorescence was found to recover spontaneously after photobleaching and displayed an intrinsic photochemistry conferring advantages over GFP-based FPs. When expressed either as a cytosolic protein or as a viral protein fusion, iLOV functioned as a superior reporter to GFP for monitoring local and systemic infections of plant RNA viruses. iLOV, therefore, offers greater utility in FP-tagging of viral gene products and represents a viable alternative where functional protein expression is limited by steric constraints or genome size.Fil: Chapman, Sean. Scottish Crop Research Institute; Reino UnidoFil: Faulkner, Christine. University of Edinburgh; Reino UnidoFil: Kaiserli, Eirini. University of Glasgow; Reino UnidoFil: Garcia-Mata, Carlos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina. University of Glasgow; Reino UnidoFil: Savenkov, Eugene I.. University of Agricultural Sciences; SueciaFil: Roberts, Alison G.. Scottish Crop Research Institute; Reino UnidoFil: Oparka, Karl J.. University of Edinburgh; Reino UnidoFil: Christie, John M.. University of Glasgow; Reino UnidoNational Academy of Sciences2008-12-16info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/138316Chapman, Sean; Faulkner, Christine; Kaiserli, Eirini; Garcia-Mata, Carlos; Savenkov, Eugene I.; et al.; The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infection; National Academy of Sciences; Proceedings of the National Academy of Sciences of The United States of America; 105; 50; 16-12-2008; 20038-200430027-8424CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.1073/pnas.0807551105info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T09:33:44Zoai:ri.conicet.gov.ar:11336/138316instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-29 09:33:44.296CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infection
title The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infection
spellingShingle The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infection
Chapman, Sean
FLUORESCENCE IMAGING
LOV DOMAIN
MOLECULAR EVOLUTION
PHOTORECEPTOR
title_short The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infection
title_full The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infection
title_fullStr The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infection
title_full_unstemmed The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infection
title_sort The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infection
dc.creator.none.fl_str_mv Chapman, Sean
Faulkner, Christine
Kaiserli, Eirini
Garcia-Mata, Carlos
Savenkov, Eugene I.
Roberts, Alison G.
Oparka, Karl J.
Christie, John M.
author Chapman, Sean
author_facet Chapman, Sean
Faulkner, Christine
Kaiserli, Eirini
Garcia-Mata, Carlos
Savenkov, Eugene I.
Roberts, Alison G.
Oparka, Karl J.
Christie, John M.
author_role author
author2 Faulkner, Christine
Kaiserli, Eirini
Garcia-Mata, Carlos
Savenkov, Eugene I.
Roberts, Alison G.
Oparka, Karl J.
Christie, John M.
author2_role author
author
author
author
author
author
author
dc.subject.none.fl_str_mv FLUORESCENCE IMAGING
LOV DOMAIN
MOLECULAR EVOLUTION
PHOTORECEPTOR
topic FLUORESCENCE IMAGING
LOV DOMAIN
MOLECULAR EVOLUTION
PHOTORECEPTOR
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Fluorescent proteins (FPs) based on green fluorescent protein (GFP) are widely used throughout cell biology to study protein dynamics, and have extensive use as reporters of virus infection and spread. However, FP-tagging of viruses is limited by the constraints of viral genome size resulting in FP loss through recombination events. To overcome this, we have engineered a smaller (10 kDa) flavinbased alternative to GFP (25 kDa) derived from the light, oxygen or voltage-sensing (LOV) domain of the plant blue light receptor, phototropin. Molecular evolution and Tobacco mosaic virus (TMV)- based expression screening produced LOV variants with improved fluorescence and photostability in planta. One variant in particular, designated iLOV, possessed photophysical properties that made it ideally suited as a reporter of subcellular protein localization in both plant and mammalian cells. Moreover, iLOV fluorescence was found to recover spontaneously after photobleaching and displayed an intrinsic photochemistry conferring advantages over GFP-based FPs. When expressed either as a cytosolic protein or as a viral protein fusion, iLOV functioned as a superior reporter to GFP for monitoring local and systemic infections of plant RNA viruses. iLOV, therefore, offers greater utility in FP-tagging of viral gene products and represents a viable alternative where functional protein expression is limited by steric constraints or genome size.
Fil: Chapman, Sean. Scottish Crop Research Institute; Reino Unido
Fil: Faulkner, Christine. University of Edinburgh; Reino Unido
Fil: Kaiserli, Eirini. University of Glasgow; Reino Unido
Fil: Garcia-Mata, Carlos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina. University of Glasgow; Reino Unido
Fil: Savenkov, Eugene I.. University of Agricultural Sciences; Suecia
Fil: Roberts, Alison G.. Scottish Crop Research Institute; Reino Unido
Fil: Oparka, Karl J.. University of Edinburgh; Reino Unido
Fil: Christie, John M.. University of Glasgow; Reino Unido
description Fluorescent proteins (FPs) based on green fluorescent protein (GFP) are widely used throughout cell biology to study protein dynamics, and have extensive use as reporters of virus infection and spread. However, FP-tagging of viruses is limited by the constraints of viral genome size resulting in FP loss through recombination events. To overcome this, we have engineered a smaller (10 kDa) flavinbased alternative to GFP (25 kDa) derived from the light, oxygen or voltage-sensing (LOV) domain of the plant blue light receptor, phototropin. Molecular evolution and Tobacco mosaic virus (TMV)- based expression screening produced LOV variants with improved fluorescence and photostability in planta. One variant in particular, designated iLOV, possessed photophysical properties that made it ideally suited as a reporter of subcellular protein localization in both plant and mammalian cells. Moreover, iLOV fluorescence was found to recover spontaneously after photobleaching and displayed an intrinsic photochemistry conferring advantages over GFP-based FPs. When expressed either as a cytosolic protein or as a viral protein fusion, iLOV functioned as a superior reporter to GFP for monitoring local and systemic infections of plant RNA viruses. iLOV, therefore, offers greater utility in FP-tagging of viral gene products and represents a viable alternative where functional protein expression is limited by steric constraints or genome size.
publishDate 2008
dc.date.none.fl_str_mv 2008-12-16
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/138316
Chapman, Sean; Faulkner, Christine; Kaiserli, Eirini; Garcia-Mata, Carlos; Savenkov, Eugene I.; et al.; The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infection; National Academy of Sciences; Proceedings of the National Academy of Sciences of The United States of America; 105; 50; 16-12-2008; 20038-20043
0027-8424
CONICET Digital
CONICET
url http://hdl.handle.net/11336/138316
identifier_str_mv Chapman, Sean; Faulkner, Christine; Kaiserli, Eirini; Garcia-Mata, Carlos; Savenkov, Eugene I.; et al.; The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infection; National Academy of Sciences; Proceedings of the National Academy of Sciences of The United States of America; 105; 50; 16-12-2008; 20038-20043
0027-8424
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.1073/pnas.0807551105
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
dc.publisher.none.fl_str_mv National Academy of Sciences
publisher.none.fl_str_mv National Academy of Sciences
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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score 13.070432

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