Bioluminescence

Uses and application in the real worldUses and application in the real world

Uses

Bioluminescence Resonance Energy Bioluminescence Resonance Energy Transfer (BRET)Transfer (BRET)

Bioluminescent molecules in gene Bioluminescent molecules in gene expressionexpression

Identification of bacterial contamination Identification of bacterial contamination in foodin food

Artistic inspirationArtistic inspiration

BRET: Bioluminescence Resonance Energy Transfer

BRET is a proximity-based BRET is a proximity-based assay where the energy assay where the energy generated by the catalytic generated by the catalytic degradation of degradation of coelenterazine by the coelenterazine by the enzyme Renilla luciferase enzyme Renilla luciferase (Rluc) (energy donor) is (Rluc) (energy donor) is transferred to a green transferred to a green fluorescent protein (GFP) fluorescent protein (GFP) acting as the energy acting as the energy acceptor. The GFP then acceptor. The GFP then emits light at its specific emits light at its specific emission wavelength. emission wavelength.

Degradation of coelenterazine by the enzyme Rluc

BRET

BRET: Bioluminescence Resonance Energy Transfer

BRET can be used to observe protein-protein BRET can be used to observe protein-protein interaction in living mammalian cells.interaction in living mammalian cells.

It is based on the non-radioactive transfer of It is based on the non-radioactive transfer of energy between a luminescent donor Rluc energy between a luminescent donor Rluc and a fluorescent acceptor (Green and a fluorescent acceptor (Green Fluorescent Protein or GFP). Fluorescent Protein or GFP).

May be used in the future to identify new May be used in the future to identify new protein complexes in human.protein complexes in human.

Monitoring of ubiquitination in living cells by BRET

Ubiquitination is descried as the rapid Ubiquitination is descried as the rapid process of post-translational modification process of post-translational modification present in many aspects of biology involving present in many aspects of biology involving a covalent attachment of ubiquitin to proteins.a covalent attachment of ubiquitin to proteins.

Monitoring of ubiquitination in ling cells with BRET

Upon degradation of Upon degradation of DeepBlueC,the DeepBlueC,the RRluc luc fuses to fuses to ββ--arrestin and arrestin and emits blue light with an emits blue light with an emission peak at 395 emission peak at 395 nm. If the nm. If the ββ-arrestin is -arrestin is ubiquitinated, a transfer ubiquitinated, a transfer of energy will occur of energy will occur between between RRluc and the luc and the GFP2 fused to the N GFP2 fused to the N terminus of ubiquitin, terminus of ubiquitin, resulting in re-emission resulting in re-emission of light with an of light with an emission peak at 510 emission peak at 510 nm.nm.

Continuous light spectrumContinuous light spectrum

Gene expression

Bioluminescence can be used to study Bioluminescence can be used to study prokaryotic gene expression inside living cellsprokaryotic gene expression inside living cells

It allows the observation of biological It allows the observation of biological processes in real time, as they happen.processes in real time, as they happen.

This technique can be used as a noninvasive This technique can be used as a noninvasive way to study protein trafficking, protein way to study protein trafficking, protein function, genetic regulatory or image bacteria, function, genetic regulatory or image bacteria, tumors and genes over a long period time.tumors and genes over a long period time.

Applications of bioluminescent imaging. (a) assessing the levels of trans-gene expression, (b) the location and extent of bacterial infection, (c) the efficiency of gene transfer and expression, and (d) the trafficking patterns of lymphocytes.

Bioluminescence in gene expression

Luciferases are light-Luciferases are light-generating enzymes generating enzymes that can be found in that can be found in bacteria, marine bacteria, marine crustaceans, fish and crustaceans, fish and insects.insects.

Luciferases are Luciferases are nontoxic and can be nontoxic and can be injected to become injected to become gene expression gene expression markers.markers.

Gene Expression in Yeast Cells

Cells fused to GFP, Cells fused to GFP, making the component making the component protein of microtubules protein of microtubules (alpha-tubulin)(alpha-tubulin)

The alpha-tubulin:GFP The alpha-tubulin:GFP fusion can be observed fusion can be observed by exciting the GFP, by exciting the GFP, causing a green light causing a green light emission.emission.

It can be used to study It can be used to study gene coding mutations gene coding mutations such as kinases.such as kinases.

Bioluminescence in Food Industry

Methods of useMethods of useATP ATP

bioluminescencebioluminescence

Bacterial Bacterial bioluminescencebioluminescence

ATP Bioluminescence

Measures the amount of ATP that is Measures the amount of ATP that is converted to photons of light by living cells.converted to photons of light by living cells.

The amount of light emitted to proportional to The amount of light emitted to proportional to the number of bacteria in a food samplethe number of bacteria in a food sample

It can be used to measure the number of It can be used to measure the number of lactic acid bacteria in a contaminated food lactic acid bacteria in a contaminated food sample.sample.

Firefly luciferase is used to detect the Firefly luciferase is used to detect the presence of ATPpresence of ATP

Luciferin and the reaction with ATP

Bacterial Bioluminescence

Can be used to test for a specific bacterial Can be used to test for a specific bacterial species or possible food born pathogen such species or possible food born pathogen such as Salmonella typhimurium.as Salmonella typhimurium.

The The luxlux gene, responsible for bacterial gene, responsible for bacterial bioluminescence, has been isolated and bioluminescence, has been isolated and cloned.cloned.

The cloned The cloned luxlux gene can be injected into a gene can be injected into a host-specific phage, which does not have the host-specific phage, which does not have the ability to express the gene.ability to express the gene.

If the phage infects a host bacterium, light If the phage infects a host bacterium, light emission results.emission results.

Detection of Bacteria

Artistic Inspirations

PoetryPoetryPhotographyPhotographyPaintingPainting

Fireflies, By Edgar FawcettI saw, one sultry night above a swamp,I saw, one sultry night above a swamp,The darkness throbbing with their golden pompThe darkness throbbing with their golden pompAnd long my dazzled sight did they entranceAnd long my dazzled sight did they entranceWith the weird chaos of their dizzy dance!With the weird chaos of their dizzy dance!Quicker than yellow leaves, when gales despoil,Quicker than yellow leaves, when gales despoil,Quivered the brilliance of their mute turmoil,Quivered the brilliance of their mute turmoil,Within whose light was intricately blentWithin whose light was intricately blentPerpetual rise, perpetual descent.Perpetual rise, perpetual descent.As though their scintillant flickerings had metAs though their scintillant flickerings had metIn the vague meshes of some airy net!In the vague meshes of some airy net!And now mysteriously I seemed to guess,And now mysteriously I seemed to guess,While watching their tumultuous loveliness,While watching their tumultuous loveliness,What fervor of deep passion strangely thrivesWhat fervor of deep passion strangely thrivesIn the warm richness of those tragic lives,In the warm richness of those tragic lives,Whose wings can never tremble but they showWhose wings can never tremble but they showThose hearts of living fire that beat below!Those hearts of living fire that beat below!

Still Photography

Artistic re-creation of the Artistic re-creation of the Bioluminescent Bay in ViequesBioluminescent Bay in Vieques

Sculpture

Montana State University-Bozeman Montana State University-Bozeman Bioglyphs projectBioglyphs project

Collaboration between art and science Collaboration between art and science during 2002 by members of the center during 2002 by members of the center for Biofilm Engineering and the MSU for Biofilm Engineering and the MSU School of artSchool of art

Lights on

Lights off

Lights on

Lights off

Involves the practice of "painting" on Involves the practice of "painting" on prepared Petri dishes with a sort of "invisible prepared Petri dishes with a sort of "invisible ink" composed of liquid medium inoculated ink" composed of liquid medium inoculated with the bacteria. with the bacteria.

The microorganisms themselves went to The microorganisms themselves went to work, multiplying on the plates and beginning work, multiplying on the plates and beginning to produce light within 24 hours.to produce light within 24 hours.

The only light available to view the art was The only light available to view the art was that produced by the bacteria themselves. that produced by the bacteria themselves. Over the five-day period, the light intensity of Over the five-day period, the light intensity of the paintings changed as the bacteria the paintings changed as the bacteria multiplied and then gradually consumed the multiplied and then gradually consumed the nutrient available.nutrient available.

Bioglyphs

Gallery, with lights on

Gallery, with lights off

Sources

Stephane Angers, Ali Salahpour, Eric Joly, Sandrine Hilairet, Stephane Angers, Ali Salahpour, Eric Joly, Sandrine Hilairet, Dan Chelsky, Michael Dennis, and Michel Bouvier. Dan Chelsky, Michael Dennis, and Michel Bouvier. “Detection of “Detection of ββ 2- 2- adrenergic receptor dimerization in living adrenergic receptor dimerization in living cells using bioluminescence resonance energy transfer cells using bioluminescence resonance energy transfer (BRET).” (BRET).” Proceedings of the National Academy of SciencesProceedings of the National Academy of Sciences 97(2000): 3684-3689. 97(2000): 3684-3689. http://www.pnas.org/cgi/content/full/97/7/3684http://www.pnas.org/cgi/content/full/97/7/3684 (5 October (5 October 2007)2007)

Claire Normand1, Stéphane Parent1, Benoit Houle, Anne Labonté, Lucie Bertrand, Mireille Caron, Mireille Legault, Stéphane Angers, Michel Bouvier, Erik C. Joly and Luc Ménard. “BRET2™: Bioluminescence Resonance Energy Transfer, a Novel Assay Technology to Examine GProtein Coupled Receptor Activation in Intact Cells.” 2002 http://las.perkinelmer.com/Content/RelatedMaterials/Posters/PSH_BRET2NovelAssayTechnology.pdf (9 October 2007). October 2007).

Saman Hosseinkhani. “Bioluminescnece.” 2007 Saman Hosseinkhani. “Bioluminescnece.” 2007 http://www.modares.ac.ir/sci/saman_h/Pages/Bioluminescencehttp://www.modares.ac.ir/sci/saman_h/Pages/Bioluminescence%20.htm%20.htm (7 October 2007). (7 October 2007).

M. W. Griffiths. “Applications of Bioluminescence in the dairy M. W. Griffiths. “Applications of Bioluminescence in the dairy Industry.” Industry.” Journal of Dairy Science Journal of Dairy Science 76 (1993): 3118-3125. 76 (1993): 3118-3125. httphttp://jds.fass.org/cgi/reprint/76/10/3118.pdf://jds.fass.org/cgi/reprint/76/10/3118.pdf (2 October 2007). (2 October 2007).

Julie Perroy, Stephanie Pontier, Pascale G Charest, Muriel Julie Perroy, Stephanie Pontier, Pascale G Charest, Muriel Aubry and Michel Bouvier. “Real-time monitoring of Aubry and Michel Bouvier. “Real-time monitoring of ubiquitination in living cells by BRET.” 1 (2004): 203-208. ubiquitination in living cells by BRET.” 1 (2004): 203-208. http://www.nature.com/nmeth/journal/v1/n3/abs/nmeth722.html;jsessihttp://www.nature.com/nmeth/journal/v1/n3/abs/nmeth722.html;jsessionid=A04B9606CE4D3B8FB5147E17BDEF094Donid=A04B9606CE4D3B8FB5147E17BDEF094D (6 October 2007). (6 October 2007).

Bouvier;s Lab. “Using BRET to study Protein-Protein interactions.” Bouvier;s Lab. “Using BRET to study Protein-Protein interactions.” June 5, 2005. June 5, 2005. http://www.mapageweb.umontreal.ca/bouvier/bret/index.htmlhttp://www.mapageweb.umontreal.ca/bouvier/bret/index.html (8 (8 October 2007).October 2007).

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Jackie Vogel. “Research.” June 3, 2005. Jackie Vogel. “Research.” June 3, 2005. http://biology.mcgill.ca/faculty/vogel/research.htmlhttp://biology.mcgill.ca/faculty/vogel/research.html (10 October (10 October 2007)2007)