Freedom for biohackers!
Fruits of lighting
Why a glowing plant will become an icon
"homemade" biotechnology of the futureAlexander Ershov, <url>
Enthusiasts donated almost half a million dollars to three young people, whom the press has already nicknamed "biohackers" – including because their work does not fall under the jurisdiction of the FDA and other government agencies that usually regulate GMOs in the United States.
"In the remote highlands of Panama, in pools guarded by guard dogs, surrounded by nets and barbed wire, swims the most expensive and studied fish in the world." This is how the Nature text begins about salmon with endogenous growth hormone production, which AquaBounty has been trying to bring to the market for 18 years.
Next year, while the fast–growing salmon is likely to swim in the Panama basin, waiting for the next approval from US government agencies, private enthusiasts without any bureaucracy will receive the seeds of the first "brainchild of systems biology" - the glowing plant Arabidopsis thaliana.
No control over the spread of the "living light bulb" is implied: according to the authors of the project, the US government has no authority to control the creation of such organisms if they are not intended for food (and this despite the fact that the plant's DNA will be equipped with not one, but a whole battery of recombinant genes). Enthusiasts who have invested in the project will be able to freely grow glowing plants, collect their seeds and even (possibly) get new glowing hybrids.
Lucifer EnzymeIn the popular press, organisms with both fluorescence and luminescence are called luminous.
Both processes occur in nature (sometimes in the same organisms) and both are used as research tools. But if fluorescence involves passive re–emission of light with a slight decrease in energy and a corresponding shift in wavelength, then luminescence is an active process during which chemical energy is converted into light.
Enzymes that convert chemical energy into light are called luciferases, and their substrates are called luciferins (both words come from the Latin lucifer – "light carrier"). Despite the same name, there is very little in common between luciferases and luciferins of, say, firefly beetles and bacteria, in addition to the actual function: they have different structures, different origins and work differently.
The most famous is the glow system of firefly beetles. During the reaction, which is catalyzed by luciferase, the ATP molecule, a universal unit of cellular energy, activates luciferin, after which oxygen is added to it (interestingly, in all organisms, despite the independent appearance of the glow system, it is somehow associated with oxidation). "Magic" occurs at the moment when CO2 is separated from the oxidized luciferin: the dye molecule is in an excited state, leaving which it emits a quantum of light.
The light produced by luciferase can have almost any shade – from blue-green to red. Here, however, there is an interesting subtlety: many coelenterates (corals, jellyfish and their relatives) have both luciferase and fluorescent proteins in their cells (for example, the notorious GFP). And usually they are not just in the same cell, but are also so closely related to each other that there is a quantum effect of excitation transfer: a photon produced by luciferase, without being emitted, is transferred to GFP, which emits it already with a changed wavelength. Therefore, invertebrates such as Renilla reniformis do not glow bluish, but green. The use of GFP makes it possible to increase the efficiency of radiation due to the fact that the fluorescent protein in comparison with luciferin of such organisms is less inclined to lose the excited state without the emission of a quantum of light. Firefly beetles, as well as luminescent bacteria, do not have proteins like GFP, and in this respect their glow system is simpler.
Scientists recognized the potential of bioluminescence for research as soon as its mechanism became known. After the advent of recombinant DNA technology, the luciferase sequence began to be "sewn" to any interesting genes and to monitor how their activation is accompanied by a glow. Pharmaceutical companies have adopted luciferase instead of the usual dye in laboratory test systems: unlike dyes, it allows you to completely get rid of extraneous "noise". Bioluminescence research, mainly in connection with various practical methods of its use, has turned into an entire branch of biotechnology with its own journals, societies, conferences and other attributes.
Nevertheless, from the point of view of biotechnology, this system is not without drawbacks. Firstly, after oxidation and the emission of a quantum of light, the spent luciferin is no longer suitable for the reaction and needs to be regenerated. Among other things, such luciferin itself inhibits the work of the enzyme. For laboratory test systems, this disadvantage does not matter much: you can simply add more substrate to the die. But if we are talking about the creation of individual luminous organisms, then the regeneration of luciferin becomes a real problem.
Secondly, if the luciferase of fireflies is encoded by only one gene and it is relatively easy to introduce it into the genome, then luciferin is an organic molecule. To synthesize it, a whole battery of genes must be inserted into the genome, which collect luciferin from pieces. Scientists have managed to achieve this goal only recently.
The first "conditionally luminous" plants obtained (.pdf) by Stephen Howell's group in 1986 (carrots and tobacco) contained only luciferase itself and did not emit light by themselves – they had to be sprayed with luciferin or added to the soil. This, by the way, can also be seen in the photographs that later became the symbol of the Glowing Plant project – the roots and vessels of tobacco glow the most, but not because luciferase works better there, but because the substrate moves along them from the soil.
The first plant capable of glowing by itself was obtained much later – only in 2010. Alexander Krichevsky and his colleagues from the universities of New York and Israel worked on it. In order to make tobacco produce its own luciferin, scientists used a block of genes from the luminescent bacteria Photobacterium leiognathi. At the same time, the genes were embedded in the genome of chloroplasts – so that they could not spread with pollen.
However, the transgenic tobacco glowed very faintly – its light is barely visible in photographs with a very long exposure. This is explained, as we already know, by the problem of luciferin regeneration, as well as by the fact that genes do not always work effectively when transferred from one organism to another. In general, such a genetic system was quite suitable for plant research (Krichevsky even registered the corresponding patent), but no one seemed to have thought about other applications until recently.
Post Tenebras LuxScientists have always considered luciferase constructs exclusively as markers capable of highlighting cells of interest to them or helping to measure the activity of the genes under study.
The very idea that such working tools can be used for something completely unrelated to research appeared in a group of students who participated in the iGEM competition, which has been organized by the Massachusetts Institute of Technology for several years. Within the framework of this competition, young scientists from different countries gather in teams, come up with and implement small projects that can somehow be attributed to the field of synthetic biology.
Participants of the iGEM-2010 contest
iGEM in many ways resembles contests of young programmers – teams are working on new, not necessarily practice-related tasks, and the result of this work, according to the organizers, should not be some global projects, but separate independent "methodological bricks" that can be used in one way or another in synthetic biology. All the projects of the competition are aimed at replenishing the so-called Standard List of biological consumables. It is assumed that this list will eventually become something like a software repository, which has been used by open source enthusiasts for many years. iGEM invented, among other things, an arsenic biosensor, a genetic system that forces bacteria to produce pigments of different colors depending on the concentration of the test substance, and even a strain of microorganisms that can be used (so far only theoretically) as a blood substitute.
It was at the iGEM competition in 2010 that nine students from the University of Cambridge decided to develop a genetic system based on luciferase from fireflies that allows creating brightly glowing organisms.
Japanese firefly Luciola cruciata, whose genes were used in the iGEM project
The students supplemented the enzymes of luciferin synthesis with an enzyme of its regeneration, optimized genes for expression in E. coli and made several more improvements. As a result, they received a strain of bacteria, a flask with which, for example, can be used instead of a lamp – such E. coli give enough light to read a book.
An iGEM contestant reads a book under the light of luminescent bacteria
Popular publications wrote about the work of students, and they themselves already dreamed of lighting roads with the help of a forest belt of glowing poplars. Interestingly, even the "green" press at that moment reacted favorably to such a fantastic idea, considering it as a way to reduce carbon dioxide emissions. But only until "fiction" began to turn into reality.
We will go to the peopleThe emergence of an audacious project, which has baffled many opponents of GMOs with its strangeness, became possible thanks to the meeting of Israeli biologist Omri Amirav-Drory and Anthony Evans, an entrepreneur with a mathematical background, imbued with futuristic ideas at Singularity University.
They were later joined by Kyle Taylor, a plant geneticist with a degree from Stanford. Being at one of the lectures where Omri talked about the potential of synthetic biology, Evans first heard about the idea of Cambridge students. And, as they say, caught fire.
There was no need to count on public or even private support for such a project, so Evans, as an IT person, decided to turn to crowdfunding on the Kickstarter website. This well-known site allows, for example, music lovers to finance the recording of albums of their favorite bands, and if the required amount cannot be collected, then the unused money is returned to users.
Evans set out to raise half a million dollars to create a glowing plant. The team promised to send the seeds of the future plant for self–cultivation to those who transferred $ 40 each for financing, and the plant itself for 150. The names of those who are ready to part with 10 thousand, scientists are going to perpetuate in the genome of the future plant (in general, we are talking about any message no longer than 30 characters).
The project aroused serious interest among Kickstarter users. In a month and a half, almost all the necessary amount was collected, and the few thousand that were not enough, Evans promised to get through the sale of T-shirts and other paraphernalia. At the same time, the plants themselves will not be sold – only those who have already invested in the project will receive them.
The future creators of the glowing plant managed to enlist the support of several famous people and organizations, among which were the founder of X Prize and Singularity University Peter Diamandis and the legendary geneticist George Church.
As an object of "highlighting", scientists have chosen a favorite model plant of geneticists – the nondescript rhesus Tal Arabidopsis thaliana. If everything goes well with him, then at the next stage biologists promise to make the rose glow.
Interestingly, no work permits for "biohackers", as the press called them, are required. The authors of the project say that under US laws, glowing plants are not subject to regulation: they are not intended for human or animal consumption, and the federal agrarian agency APHIS (Animal and Plant Health Inspection Service) is only interested in the method of introducing transgenes. If the genes of the luciferase system are introduced into the plant by a method that does not use pathogens (originally it was supposed to use the conditionally pathogenic Agrobacterium for this), APHIS will not be able to interfere with the work of Evans and his colleagues.
A set for self-transformation of plants from Glowing Plant
Evans and his associates promise to send the project participants a set for self–transformation of plants, but whether they will be able to do it remains a big question. The bacteria that are used for such a simple home transformation are conditionally pathogenic, so their spread may fall under legal restrictions.
It is not surprising that such freedom caused some "greens" a storm of indignation. They have already called the creation of glowing plants a loophole in US laws, which "irresponsible hackers" can use as they please. An article calling for urgently patching up a gap in the laws appeared, in addition to the profile eco-activist press, in The Guardian. The Canadian organization ETC Group behaved most actively – it tried to organize a public campaign designed to put pressure on Kickstarter in order not to give funding to "biohackers".
Omphalotus olearius is one of many naturally luminescent fungi,
which do not cause protests by eco-activists.
It is difficult to say whether Evans and his colleagues will be able to overcome the resistance and deliver the seeds to the enthusiasts who supported the project on time. So far it seems that they have a much better chance of this than AquaBounty with their long-suffering salmon. After all, "biohackers", unlike AquaBounty, Monsanto and other biotech companies, will have to fight not with the American bureaucracy in the form of the FDA, but with public opinion and eco-activists, who, fortunately or unfortunately, are not so powerful yet.
Portal "Eternal youth" http://vechnayamolodost.ru24.06.2013