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An endogenous green fluorescent protein-photoprotein pair in Clytia hemisphaerica eggs shows co-targeting to mitochondria and efficient bioluminescence energy transfer.

Fourrage C, Swann K, Gonzalez Garcia JR, Campbell AK, Houliston E - Open Biol (2014)

Bottom Line: Green fluorescent proteins (GFPs) and calcium-activated photoproteins of the aequorin/clytin family, now widely used as research tools, were originally isolated from the hydrozoan jellyfish Aequora victoria.Using the hydrozoan Clytia hemisphaerica, we characterized differential expression of three clytin and four GFP genes in distinct tissues at larva, medusa and polyp stages, corresponding to the major in vivo sites of bioluminescence (medusa tentacles and eggs) and fluorescence (these sites plus medusa manubrium, gonad and larval ectoderms).Remarkably, the clytin2 and GFP2 proteins, co-expressed in eggs, show particularly efficient BRET and co-localize to mitochondria, owing to parallel acquisition by the two genes of mitochondrial targeting sequences during hydrozoan evolution.

View Article: PubMed Central - PubMed

Affiliation: Sorbonne Universités, UPMC Univ Paris 06, Laboratoire de Biologie du Développement de Villefranche-sur-mer (LBDV), Observatoire Océanologique, 06230 Villefranche-sur-mer, France.

ABSTRACT
Green fluorescent proteins (GFPs) and calcium-activated photoproteins of the aequorin/clytin family, now widely used as research tools, were originally isolated from the hydrozoan jellyfish Aequora victoria. It is known that bioluminescence resonance energy transfer (BRET) is possible between these proteins to generate flashes of green light, but the native function and significance of this phenomenon is unclear. Using the hydrozoan Clytia hemisphaerica, we characterized differential expression of three clytin and four GFP genes in distinct tissues at larva, medusa and polyp stages, corresponding to the major in vivo sites of bioluminescence (medusa tentacles and eggs) and fluorescence (these sites plus medusa manubrium, gonad and larval ectoderms). Potential physiological functions at these sites include UV protection of stem cells for fluorescence alone, and prey attraction and camouflaging counter-illumination for bioluminescence. Remarkably, the clytin2 and GFP2 proteins, co-expressed in eggs, show particularly efficient BRET and co-localize to mitochondria, owing to parallel acquisition by the two genes of mitochondrial targeting sequences during hydrozoan evolution. Overall, our results indicate that endogenous GFPs and photoproteins can play diverse roles even within one species and provide a striking and novel example of protein coevolution, which could have facilitated efficient or brighter BRET flashes through mitochondrial compartmentalization.

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Related in: MedlinePlus

Relative expression of CheGFP and clytin genes evaluated by Q-PCR. (a) Q-PCR analysis showing relative expression (% of total) of each GFP gene. (b) Schematic summary of GFP differential expression in Clytia. (c) Relative expression (% of total) of each clytin gene measured by Q-PCR. (d) Schematic summary of clytin differential expression in Clytia. All Q-PCR primer sequences are provided in the electronic supplementary material, table S1. Annotations for all panels: Pl, planulae; Um, bell; M, manubrium; G, gonad; Ge, gonad ectoderm; Tb, tentacle bulb; E, eggs.
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RSOB130206F4: Relative expression of CheGFP and clytin genes evaluated by Q-PCR. (a) Q-PCR analysis showing relative expression (% of total) of each GFP gene. (b) Schematic summary of GFP differential expression in Clytia. (c) Relative expression (% of total) of each clytin gene measured by Q-PCR. (d) Schematic summary of clytin differential expression in Clytia. All Q-PCR primer sequences are provided in the electronic supplementary material, table S1. Annotations for all panels: Pl, planulae; Um, bell; M, manubrium; G, gonad; Ge, gonad ectoderm; Tb, tentacle bulb; E, eggs.

Mentions: Fluorescence in C. hemisphaerica. Green fluorescence observed upon excitation with blue light under a stereomicroscope (a), fluorescence microscope (b,f) or confocal microscope with 488 nm laser excitation (c,d,e,g), superimposed on transmitted light images in (d,e,g). (a) Adult medusa immobilized in agar (courtesy of A. Amiel); M, manubrium; G, gonad; Tb, tentacle bulb. (b) Bell margin with tentacle bulb (baby medusa), (c) manubrium (baby medusa), (d) tentacle bulb, (e) tentacle, (f) planula larva (2 days post fertilization), (g) stolon. For a guide to medusa organization, see figure 4. All scale bars 100 µm.


An endogenous green fluorescent protein-photoprotein pair in Clytia hemisphaerica eggs shows co-targeting to mitochondria and efficient bioluminescence energy transfer.

Fourrage C, Swann K, Gonzalez Garcia JR, Campbell AK, Houliston E - Open Biol (2014)

Relative expression of CheGFP and clytin genes evaluated by Q-PCR. (a) Q-PCR analysis showing relative expression (% of total) of each GFP gene. (b) Schematic summary of GFP differential expression in Clytia. (c) Relative expression (% of total) of each clytin gene measured by Q-PCR. (d) Schematic summary of clytin differential expression in Clytia. All Q-PCR primer sequences are provided in the electronic supplementary material, table S1. Annotations for all panels: Pl, planulae; Um, bell; M, manubrium; G, gonad; Ge, gonad ectoderm; Tb, tentacle bulb; E, eggs.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4043110&req=5

RSOB130206F4: Relative expression of CheGFP and clytin genes evaluated by Q-PCR. (a) Q-PCR analysis showing relative expression (% of total) of each GFP gene. (b) Schematic summary of GFP differential expression in Clytia. (c) Relative expression (% of total) of each clytin gene measured by Q-PCR. (d) Schematic summary of clytin differential expression in Clytia. All Q-PCR primer sequences are provided in the electronic supplementary material, table S1. Annotations for all panels: Pl, planulae; Um, bell; M, manubrium; G, gonad; Ge, gonad ectoderm; Tb, tentacle bulb; E, eggs.
Mentions: Fluorescence in C. hemisphaerica. Green fluorescence observed upon excitation with blue light under a stereomicroscope (a), fluorescence microscope (b,f) or confocal microscope with 488 nm laser excitation (c,d,e,g), superimposed on transmitted light images in (d,e,g). (a) Adult medusa immobilized in agar (courtesy of A. Amiel); M, manubrium; G, gonad; Tb, tentacle bulb. (b) Bell margin with tentacle bulb (baby medusa), (c) manubrium (baby medusa), (d) tentacle bulb, (e) tentacle, (f) planula larva (2 days post fertilization), (g) stolon. For a guide to medusa organization, see figure 4. All scale bars 100 µm.

Bottom Line: Green fluorescent proteins (GFPs) and calcium-activated photoproteins of the aequorin/clytin family, now widely used as research tools, were originally isolated from the hydrozoan jellyfish Aequora victoria.Using the hydrozoan Clytia hemisphaerica, we characterized differential expression of three clytin and four GFP genes in distinct tissues at larva, medusa and polyp stages, corresponding to the major in vivo sites of bioluminescence (medusa tentacles and eggs) and fluorescence (these sites plus medusa manubrium, gonad and larval ectoderms).Remarkably, the clytin2 and GFP2 proteins, co-expressed in eggs, show particularly efficient BRET and co-localize to mitochondria, owing to parallel acquisition by the two genes of mitochondrial targeting sequences during hydrozoan evolution.

View Article: PubMed Central - PubMed

Affiliation: Sorbonne Universités, UPMC Univ Paris 06, Laboratoire de Biologie du Développement de Villefranche-sur-mer (LBDV), Observatoire Océanologique, 06230 Villefranche-sur-mer, France.

ABSTRACT
Green fluorescent proteins (GFPs) and calcium-activated photoproteins of the aequorin/clytin family, now widely used as research tools, were originally isolated from the hydrozoan jellyfish Aequora victoria. It is known that bioluminescence resonance energy transfer (BRET) is possible between these proteins to generate flashes of green light, but the native function and significance of this phenomenon is unclear. Using the hydrozoan Clytia hemisphaerica, we characterized differential expression of three clytin and four GFP genes in distinct tissues at larva, medusa and polyp stages, corresponding to the major in vivo sites of bioluminescence (medusa tentacles and eggs) and fluorescence (these sites plus medusa manubrium, gonad and larval ectoderms). Potential physiological functions at these sites include UV protection of stem cells for fluorescence alone, and prey attraction and camouflaging counter-illumination for bioluminescence. Remarkably, the clytin2 and GFP2 proteins, co-expressed in eggs, show particularly efficient BRET and co-localize to mitochondria, owing to parallel acquisition by the two genes of mitochondrial targeting sequences during hydrozoan evolution. Overall, our results indicate that endogenous GFPs and photoproteins can play diverse roles even within one species and provide a striking and novel example of protein coevolution, which could have facilitated efficient or brighter BRET flashes through mitochondrial compartmentalization.

Show MeSH
Related in: MedlinePlus