Plants with genetically encoded autoluminescence.

Autoluminescent plants engineered to express a bacterial bioluminescence gene cluster in plastids have not been widely adopted because of low light output. We engineered tobacco plants with a fungal bioluminescence system that converts caffeic acid (present in all plants) into luciferin and report self-sustained luminescence that is visible to the naked eye. Our findings could underpin development of a suite of imaging tools for plants.

Genetically encodable bioluminescent system from fungi.

Bioluminescence is found across the entire tree of life, conferring a spectacular set of visually oriented functions from attracting mates to scaring off predators. Half a dozen different luciferins, molecules that emit light when enzymatically oxidized, are known. However, just one biochemical pathway for luciferin biosynthesis has been described in full, which is found only in bacteria. Here, we report identification of the fungal luciferase and three other key enzymes that together form the biosynthetic cycle of the fungal luciferin from caffeic acid, a simple and widespread metabolite. Introduction of the identified genes into the genome of the yeast Pichia pastoris along with caffeic acid biosynthesis genes resulted in a strain that is autoluminescent in standard media. We analyzed evolution of the enzymes of the luciferin biosynthesis cycle and found that fungal bioluminescence emerged through a series of events that included two independent gene duplications. The retention of the duplicated enzymes of the luciferin pathway in nonluminescent fungi shows that the gene duplication was followed by functional sequence divergence of enzymes of at least one gene in the biosynthetic pathway and suggests that the evolution of fungal bioluminescence proceeded through several closely related stepping stone nonluminescent biochemical reactions with adaptive roles. The availability of a complete eukaryotic luciferin biosynthesis pathway provides several applications in biomedicine and bioengineering.

Progress in the Study of Bioluminescent Earthworms.

Even though bioluminescent oligochaetes rarely catch people's eyes due to their secretive lifestyle, glowing earthworms sighting reports have come from different areas on all continents except Antarctica. A major breakthrough in the research of earthworm bioluminescence occurred in the 1960s with the studies of the North American Diplocardia longa. Comparative studies conducted on 13 earthworm species belonging to six genera showed that N-isovaleryl-3-aminopropanal (Diplocardia luciferin) is the common substrate for bioluminescence in all examined species, while luciferases appeared to be responsible for the color of bioluminescence. The second momentous change in the situation has occurred with the discovery in Siberia (Russia) of two unknown luminous enchytraeids. The two bioluminescent systems belong to different types, have different spectral characteristics and localization, and different temperature and pH optima. They are unique, and this fact is confirmed by the negative results of all possible cross-reactions. The bioluminescent system of Henlea sp. comprises four essential components: luciferase, luciferin, oxygen and calcium ion. For Friderica heliota, the luminescent reaction requires five components: luciferase, luciferin, ATP, magnesium ion and oxygen. Along with luciferin, more than a dozen analogues were isolated from worm biomass. These novel peptide-like natural compounds represent an unprecedented chemistry found in terrestrial organisms.

1001 lights: luciferins, luciferases, their mechanisms of action and applications in chemical analysis, biology and medicine.

Bioluminescence (BL) is a spectacular phenomenon involving light emission by live organisms. It is caused by the oxidation of a small organic molecule, luciferin, with molecular oxygen, which is catalysed by the enzyme luciferase. In nature, there are approximately 30 different BL systems, of which only 9 have been studied to various degrees in terms of their reaction mechanisms. A vast range of in vitro and in vivo analytical techniques have been developed based on BL, including tests for different analytes, immunoassays, gene expression assays, drug screening, bioimaging of live organisms, cancer studies, the investigation of infectious diseases and environmental monitoring. This review aims to cover the major existing applications for bioluminescence in the context of the diversity of luciferases and their substrates, luciferins. Particularly, the properties and applications of d-luciferin, coelenterazine, bacterial, Cypridina and dinoflagellate luciferins and their analogues along with their corresponding luciferases are described. Finally, four other rarely studied bioluminescent systems (those of limpet Latia, earthworms Diplocardia and Fridericia and higher fungi), which are promising for future use, are also discussed.

A Tale Of Two Luciferins: Fungal and Earthworm New Bioluminescent Systems.

Bioluminescence, the ability of a living organism to produce light through a chemical reaction, is one of Nature's most amazing phenomena widely spread among marine and terrestrial species. There are various different mechanisms underlying the emission of "cold light", but all involve a small molecule, luciferin, that provides energy for light-generation upon oxidation, and a protein, luciferase, that catalyzes the reaction. Different species often use different proteins and substrates in the process, which suggests that the ability to produce light evolved independently several times throughout evolution. Currently, it is estimated that there are more than 30 different mechanisms of bioluminescence. Even though the chemical foundation underlying the bioluminescence phenomenon is by now generally understood, only a handful of luciferins have been isolated and characterized. Today, the known bioluminescence reactions are used as indispensable analytical tools in various fields of science and technology. A pressing need for new bioluminescent analytical techniques with a wider range of practical applications stimulates the search and chemical studies of new bioluminescent systems. In the past few years two such systems were unraveled: those of the earthworms Fridericia heliota and the higher fungi. The luciferins of these two systems do not share structural similarity with the previously known ones. This Account will survey structure elucidation of the novel luciferins and identification of their mechanisms of action. Fridericia luciferin is a key component of a novel ATP-dependent bioluminescence system. Structural studies were performed on 0.005 mg of natural substance and revealed its unusual extensively modified peptidic nature. Elucidation of Fridericia oxyluciferin revealed that oxidative decarboxylation of a lysine fragment of luciferin supplies energy for light generation, while a fluorescent CompX moiety remains intact and serves as a light emitter. Along with luciferin, a number of its natural analogs were found in the extracts of worm biomass. They occurred to be highly unusual modified peptides comprising a set of amino acids, including threonine, aminobutyric acid, homoarginine, unsymmetrical N,N-dimethylarginine and extensively modified tyrosine. These natural compounds represent a unique peptide chemistry found in terrestrial animals and raise novel questions concerning their biosynthetic origin. Also in this Account we discuss identification of the luciferin of higher fungi 3-hydroxyhispidin which is biosynthesized by oxidation of the precursor hispidin, a known fungal and plant secondary metabolite. Furthermore, it was shown that 3-hydroxyhispidin leads to bioluminescence in extracts from four diverse genera of luminous fungi, thus suggesting a common biochemical mechanism for fungal bioluminescence.

Novel peptide chemistry in terrestrial animals: natural luciferin analogues from the bioluminescent earthworm Fridericia heliota.

We report isolation and structure elucidation of AsLn5, AsLn7, AsLn11 and AsLn12: novel luciferin analogs from the bioluminescent earthworm Fridericia heliota. They were found to be highly unusual modified peptides, comprising either of the two tyrosine-derived chromophores, CompX or CompY and a set of amino acids, including threonine, gamma-aminobutyric acid, homoarginine, and unsymmetrical N,N-dimethylarginine. These natural compounds represent a unique peptide chemistry found in terrestrial animals and rise novel questions concerning their biosynthetic origin.

A novel type of luciferin from the Siberian luminous earthworm Fridericia heliota: structure elucidation by spectral studies and total synthesis.

The structure elucidation and synthesis of the luciferin from the recently discovered luminous earthworm Fridericia heliota is reported. This luciferin is a key component of a novel ATP-dependent bioluminescence system. UV, fluorescence, NMR, and HRMS spectroscopy studies were performed on 0.005 mg of the isolated substance and revealed four isomeric structures that conform to spectral data. These isomers were chemically synthesized and one of them was found to produce light when reacted with a protein extract from F. heliota. The novel luciferin was found to have an unusual extensively modified peptidic nature, thus implying an unprecedented mechanism of action.