title: Salicylic acid interferes with GFP fluorescence in vivo.
abstract: Fluorescent proteins have become essential tools for cell biologists. They are routinely used by plant biologists for protein and promoter fusions to infer protein localization, tissue-specific expression and protein abundance. When studying the effects of biotic stress on chromatin, we unexpectedly observed a decrease in GFP signal intensity upon salicylic acid (SA) treatment in Arabidopsis lines expressing histone H1-GFP fusions. This GFP signal decrease was dependent on SA concentration. The effect was not specific to the linker histone H1-GFP fusion but was also observed for the nucleosomal histone H2A-GFP fusion. This result prompted us to investigate a collection of fusion proteins, which included different promoters, subcellular localizations and fluorophores. In all cases, fluorescence signals declined strongly or disappeared after SA application. No changes were detected in GFP-fusion protein abundance when fluorescence signals were lost indicating that SA does not interfere with protein stability but GFP fluorescence. In vitro experiments showed that SA caused GFP fluorescence reduction only in vivo but not in vitro, suggesting that SA requires cellular components to cause fluorescence reduction. Together, we conclude that SA can interfere with the fluorescence of various GFP-derived reporter constructs in vivo. Assays that measure relocation or turnover of GFP-tagged proteins upon SA treatment should therefore be evaluated with caution.
authors.affiliation: Swedish University of Agricultural Sciences, Department of Plant Biology and Linnean Center for Plant Biology, PO Box 7080, SE-75007 Uppsala, Sweden.
authors.forename: Jennifer
authors.initials: J
authors.lastname: de Jonge
authors.affiliation: Swedish University of Agricultural Sciences, Department of Plant Biology and Linnean Center for Plant Biology, PO Box 7080, SE-75007 Uppsala, Sweden.
authors.forename: Daniel
authors.initials: D
authors.lastname: Hofius
authors.forename: Lars
authors.initials: L
authors.lastname: Hennig
mesh_terms: D017360:Arabidopsis / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics* / Q000378:metabolism; D005453:Fluorescence; D049452:Green Fluorescent Proteins / Q000235:genetics* / Q000378:metabolism; D006657:Histones / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D020156:Salicylic Acid / Q000008:administration & dosage / Q000378:metabolism*
chemical_list: D029681:Arabidopsis Proteins; D006657:Histones; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D020156:Salicylic Acid
keywords: Arabidopsis; GFP fusion proteins; fluorescence microscopy; fluorescent protein; histones; salicylic acid.