A bright monomeric green fluorescent protein derived from Branchiostoma lanceolatum

Shaner, Nathan C.; Lambert, G.; Chammas, Andrew; Ni, Yuhui; Cranfill, Paula J.; Baird, Michelle A.; Sell, Brittney R.; Allen, John R.; Day, Richard N.; Israelsson, Maria; Davidson, Michael W.; Wang, Jiwu

doi:10.1038/nmeth.2413

Cited by 1,120 publications

(1,122 citation statements)

References 28 publications

(45 reference statements)

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“…Each FP also has a characteristic maturation period (i.e., the time it takes for the newly synthesized FP polypeptide to fold into its mature fluorescent conformation). Although rapid maturation is beneficial for most applications (the dLanYFP derivative mNeonGreen requires <1 min whereas GFP requires~25 min; Shaner et al 2013), slow-folding RFPs requiring several hours (Baird et al 2000) support lineage analysis of organelle inheritance over successive rounds of cell division (Grallert et al 2004;Lam et al 2012). Maximum projections from 21 z-plane images of cam1-gfp (left panels), cam1-yfp (center panels), and cam1-mCherry (right panels) cells.…”

Section: Fluorescent Proteinsmentioning

confidence: 99%

“…Although GFP has long been an excellent FP for most imaging applications, its position will be continually challenged making it important to consult members of the fission yeast community involved in imaging (e.g., via Pomblist: http://listserver.ebi.ac.uk/mailman/listinfo/pombelist) to establish which probes are best suited for fission yeast imaging. At the time of writing, the similar absorption and emission spectra and rapid folding yet threefold brighter signal suggest that mNeonGreen will likely be the first to eclipse GFP (Shaner et al 2013). …”

Section: Fluorescent Proteinsmentioning

confidence: 99%

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Live Cell Imaging in Fission Yeast

Mulvihill

2017

Cold Spring Harb Protoc

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Live cell imaging complements the array of biochemical and molecular genetic approaches to provide a comprehensive insight into functional dependencies and molecular interactions in fission yeast. Fluorescent proteins and vital dyes reveal dynamic changes in the spatial distribution of organelles and the proteome and how each alters in response to changes in environmental and genetic composition. This introduction discusses key issues and basic image analysis for live cell imaging of fission yeast.

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Section: Fluorescent Proteinsmentioning

confidence: 99%

Section: Fluorescent Proteinsmentioning

confidence: 99%

Live Cell Imaging in Fission Yeast

Mulvihill

2017

Cold Spring Harb Protoc

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“…At this time, we recommend superfolder GFP [56, 57], secBFP2 [47], and FusionRed [58] for protein fusions. For transcriptional reporters, mNeonGreen matures rapidly and produces an intense signal [59] and TagRFP has similar properties for a red reporter [60]. …”

Section: Approaches For Imaging Er Stress and Upr Activity In Livinmentioning

confidence: 99%

“…FPs are generally not bright enough to image at low expression levels in live cells [106]. Some newer brighter FPs, such as mNeonGreen [59] have been reported and suggest that it may be possible to image low expressed proteins at physiologic levels in cells.…”

Section: Perspectivementioning

confidence: 99%

Approaches to imaging unfolded secretory protein stress in living cells

Lajoie

Fazio

Snapp

2014

Endoplasmic Reticulum Stress in Diseases

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The endoplasmic reticulum (ER) is the point of entry of proteins into the secretory pathway. Nascent peptides interact with the ER quality control machinery that ensures correct folding of the nascent proteins. Failure to properly fold proteins can lead to loss of protein function and cytotoxic aggregation of misfolded proteins that can lead to cell death. To cope with increases in the ER unfolded secretory protein burden, cells have evolved the Unfolded Protein Response (UPR). The UPR is the primary signaling pathway that monitors the state of the ER folding environment. When the unfolded protein burden overwhelms the capacity of the ER quality control machinery, a state termed ER stress, sensor proteins detect accumulation of misfolded peptides and trigger the UPR transcriptional response. The UPR, which is conserved from yeast to mammals, consists of an ensemble of complex signaling pathways that aims at adapting the ER to the new misfolded protein load. To determine how different factors impact the ER folding environment, various tools and assays have been developed. In this review, we discuss recent advances in live cell imaging reporters and model systems that enable researchers to monitor changes in the unfolded secretory protein burden and activation of the UPR and its associated signaling pathways.

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“…), and could be successfully evolved into the monomeric green fluorescent protein mNeonGreen, which retains 65% of the brightness of the parent protein (Shaner et al, 2013). Monomerization of lanYFP was achieved in a series of rounds of mutagenesis based on rational design and directed evolution.…”

mentioning

confidence: 99%

Structural analysis of the bright monomeric yellow-green fluorescent protein mNeonGreen obtained by directed evolution

Clavel

Gotthard

Stetten

et al. 2016

Acta Cryst Sect D Struct Biol

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Until recently, genes coding for homologues of the autofluorescent protein GFP had only been identified in marine organisms from the phyla Cnidaria and Arthropoda. New fluorescent-protein genes have now been found in the phylum Chordata, coding for particularly bright oligomeric fluorescent proteins such as the tetrameric yellow fluorescent protein lanYFP from Branchiostoma lanceolatum. A successful monomerization attempt led to the development of the bright yellow-green fluorescent protein mNeonGreen. The structures of lanYFP and mNeonGreen have been determined and compared in order to rationalize the directed evolution process leading from a bright, tetrameric to a still bright, monomeric fluorescent protein. An unusual discolouration of crystals of mNeonGreen was observed after X-ray data collection, which was investigated using a combination of X-ray crystallography and UV-visible absorption and Raman spectroscopies, revealing the effects of specific radiation damage in the chromophore cavity. It is shown that X-rays rapidly lead to the protonation of the phenolate O atom of the chromophore and to the loss of its planarity at the methylene bridge.

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A bright monomeric green fluorescent protein derived from Branchiostoma lanceolatum

Cited by 1,120 publications

References 28 publications

Live Cell Imaging in Fission Yeast

Live Cell Imaging in Fission Yeast

Approaches to imaging unfolded secretory protein stress in living cells

Structural analysis of the bright monomeric yellow-green fluorescent protein mNeonGreen obtained by directed evolution

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