Publish date: 02.05.2018
Synthetic biology involves the modification and design of whole organisms to solve various problems, for example, the production of biosensors and various useful compounds in bioreactors, the restoration of certain ecological niches (bioremediation), and so on. Most often, genetically modified microorganisms (GMMs) are used for these purposes because microorganisms are much simpler than multicellular organisms and consequently are easier to transform.
In the course of development synthetic biology is progressing from theoretical ideas and laboratory experiments to practical applications. However, there remain fears that artificial microbes can cause undesirable consequences when they enter the environment or get into the unintended ecological niche.
It should be noted that the safety principles of genetic engineering and the corresponding recommendations were adopted in 1975 at a conference in Asilomare (California, USA). According to these recommendations, engineered organisms are designed in such a way that their survival in the environment is as short as possible (for example, weeks, not years). The usual physical barriers (containers, filtration systems, safety equipment) that prevent the entry of the GMMs into unintended ecological niches also should not be underestimated. In addition, artificially created organisms due to modifications are usually not able to survive in nature.
However at present genetic engineering allows the creation of more adapted for survival organisms, and therefore the safety measures are also being revised. Modern approaches to the safety of GMMs are discussed in the review: Wright O, Stan G-B and Ellis T. Building-in biosafety for synthetic biology. Microbiology (2013), 159, 1221-1235.
One of the most important problems is the horizontal gene transfer, a phenomenon common among microorganisms. It occurs through plasmids (conjugation), bacteriophages (transduction), or by transformation. For example, when constructing genetically engineered microbes, plasmids containing antibiotic resistance genes are often used as selective markers. The entry of such genes into pathogenic microbes is extremely undesirable.
The second possible problem is the entry of modified microbes into the environment and into animal’s and human’s bodies, where they can disrupt the natural balance. In order to prevent these incidents, scientists developed several approaches. One of them involves the use of toxin-antitoxin pairs. In this case, the synthesis of the toxin is encoded in the plasmid, and the synthesis of the antitoxin -in the chromosome. Thus, if this plasmid enters another microorganism, it immediately dies, as it starts to synthesize a toxin, but there is no antitoxin in this bacterium. Another approach is to use auxotrophy. In this case, one of the necessary genes is deactivated in the chromosome such as a gene responsible for the synthesis of one of the essential amino acids, and the same gene is introduced into the plasmid. The survival of such an organism is possible only when the plasmid is conserved within the bacteria. If the horizontal gene transfer does occur, and the plasmid has got into another organism, it will not be retained for a long time because it represents an additional burden on the cell.
Another approach to combating horizontal gene transfer is to transfer part of the genetic code for proteins involved in plasmid replication to the chromosome in such a way that this plasmid cannot be replicated in any other microorganism, except for a specially designed host microbe.
Often a combination of several methods is used.
All these systems, however, do not guarantee the absolute safety. Modified bacteria can overcome these protection mechanisms as a result of random mutations or, in the case of auxotrophs, get the missing nutrient from the environment, and also borrow it from other bacteria.
In 2016, scientists created new, multi-level control systems with increased reliability. These are the so-called self-destruct buttons “Deadman” and “Passcode” (Chan CTY, Lee JW, Cameron DE, Bashor CJ and Collins JJ. “Deadman” and “Passcode” microbial kill switches for bacterial containment. Nature chemical biology (2016), 12, 82-86).
In “Deadman” systems, sensor proteins detect certain substances – signaling molecules – in the environment and, if present, trigger the synthesis of a toxin that kills microorganisms and/or stops the synthesis of the antitoxin. The systems are duplicated for better reliability. For example, there may be two or more different toxins (each paired with its signal molecule), so that a system failure due to one random mutation is impossible. In addition, it is possible to directly induce toxin synthesis bypassing these protein-sensors.
The “Passcode” system adds one more level of protection. Hybrid transcription factors that are used here can recognize several signal molecules in the environment simultaneously, and only if all these molecules are present will the organism be able to grow. The system is modular in nature, so if necessary it can be easily reprogrammed. For example, the GMM will grow only if galactose and glucose are present in the medium simultaneously. At the same time only the laboratory or factory staff know the set of specific signal molecules necessary for the growth of microorganisms (hence the name “Passcode”). Thus, this system protects not only from getting of GMMs into the environment, but also from their “pirate” use (patent infringement, industrial espionage).
In order to track the possible entry of genetically modified organisms into the environment, the so-called DNA barcodes are used. These are synthetic DNA sequences that are inserted into the genome of modified organisms and can be read by sequencing of the samples from the environment. The same methods are used to prevent industrial espionage.
Most of the above mentioned security systems are already used in the process of construction of the GMMs that are already available on the market.
A new promising approach is the development of the so-called semantic barrier by introducing changes in the genetic code of bacteria. In the process of the organisms’ design and engineering, scientists use alternative stop codons, nucleotide quadruplets instead of triplets and even new synthetic nucleotides that do not occur in nature, in order to encode the same proteins. Such DNA could not function when transferred to another organism (this can be compared to how, for example, it is difficult for people in a foreign country without knowing the language, so the barrier is called semantic). In that case, the organisms themselves would be wholly dependent on artificial metabolites, and, therefore, could not grow outside the bioreactor. Though the first successful results have already been published, these methods are still far from practical application.
Synthetic biology has tremendous potential. In order for it to be fully implemented, new security systems should be developed and the already existing systems should be improved. A different set of security measures is needed depending on which organism is designed, what is its purpose and how it is supposed to be used. Today it is safe to say that scientists succeeded in developing systems with high levels of safety for the artificially created organisms, and research in this direction continues.
journalist, founder of the popular science YouTube channel ‘Ratiomania’