Molecular Biotechnology: from the Dynamics of Biological Systems to Applications

In the elapsed program period, the program group ''Molecular biotechnology'' has established itself internationally particularly in the fields of synthetic biology and immunology. In the upcoming program period we plan to further intertwine those two research fields. The major focuses of the program will be directed towards: - designed bionanostructures based on biological molecules, - the design of regulatory networks in cells, particularly in mammalian cells, - understanding innate immune responses connected with disease, - the combination of synthetic biology and immunology for the preparation of new therapeutic interventions and other possible uses of synthetic biology tools. Proteins are biopolymers, which perform the majority of functions in the cell. Recently, we published a very innovative principle for modular folding of polypeptides (Gradišar et al., Nat.Chem.Biol. 2013), which represents an important breakthrough in the design of bionanostructures and the foundation for our further research in this field. Within this program proposal, we plan to design modular proteins with the capability of self-assembling in vivo. Further, we plan to decorate our nanostructures with selected protein domains in order to be tested as novel vaccines. Our goal is also to prepare more complex designed protein polyhedra and control their assembly and disassembly. We plan to prepare lattices composed of different polypeptide units. The final goal of this part is to translate our achievements into concrete applications in the fields of medicine, sensors and biosynthesis. In parallel to polypeptide-based materials we plan to develop nanostructures based on DNA and polypeptide-DNA hybrid nanostructures. Cells can be modified for therapeutic purposes by the manipulation of their response to external triggers and intracellular processes. In this program proposal we plan to upgrade our recent achievements in cellular logics (Gaber et al., Nat.Chem.Biol., 2014). In addition, we plan to develop synthetic biology tools based on CRISPR/Cas and TALE to perform logic operations and genome modifications, which represent the bottleneck of the advanced use of synthetic biology in mammalian cell. Innate immune response is extremely important for human health, and is particularly deeply involved in chronic inflammatory diseases. During the next program period we plan to define unknown mechanisms of activation of membrane and cytosolic innate immunity receptors, TLRs and NLRs and the connected signaling pathways. We plan to investigate the relevance of our discoveries  regarding human health in close collaboration with physicians on samples of patients suffering from chronic diseases.

The proposed research program will respond to important issues in the fields of life sciences, especially in medically important processes. In addition, we intend to prepare new synthetic biology tools in both bionanomaterials and cellular logic fields. Such tools will be very useful for researchers in the field of synthetic biology, as well as the broader biotechnology and life sciences in general. Our recent discovery of a new type of modular protein folds and design of cellular logic in mammalian cells using modular DNA-binding proteins, have attracted much attention in the world. That resulted in invitations to lecture at two Gordon Research Conferences in 2014 and to an additional one in 2015. Our publication about the preparation of the tetrahedron was commented on in the journals Nature, Science, Nature Biotechnology, and C&E News, therefore we expect that the upgrade of this successful research will be even more notable. With the preparation of the tetrahedron from one polypeptide chain we opened a new direction of research in the field of bionanotechnology. This discovery is inspired by DNA nanotechnology, but due to the characteristics of the polypeptides and the possibility of preparation of cell factories, has enormous potential for actual applications and for industrial use. The proposed objectives of the program will be a big step towards moving the frontiers of technology and to expand the research groups active in this area. The knowledge about the molecular recognition mechanisms in the immune system is important for the understanding of physiological processes in cells and organisms and essential for drug design. Knowledge about the molecular mechanism of activation of TLRs and NLRs is important for understanding of the functioning of innate immunity. We can expect high-profile publications in reputable scientific journals in the mentioned fields. Understanding of the activation mechanism of innate immunity receptors is one of the fundamental problems of immunology. The implementation of this knowledge in therapeutic applications is very important. Due to the broad role of TLRs in sterile inflammatory processes the relevance of these studies ranges from the fields of autoimmunity, cancer, and neuropathic pain to many other diseases. The innovative combination of synthetic biology and immunology has a great potential to become a fine example of the relevance of synthetic biology for health.

Based on the high international recognition of the scientific achievements and by providing training to highly qualified personnel, the research program Molecular biotechnology has a substantial direct importance for the society. In the recent years, the pharmaceutical and biotech industry expressed a strong interest for employing our researchers, which is evident from the employment of 9 researchers who have been trained in our program group. Moreover, tens of students that obtained their training within the iGEM project continue their research career in other program groups as well as at top universities abroad. These students represent a bridge between the Slovenian science and science abroad and are potential staff that can return to Slovenia after completion of education. Since 2006 our program group contributed significantly to the promotion of life sciences and science in general to a wider audience through exceptional successes in competitions of research projects. This promoted Slovenia abroad as a country with excellent science and education. Hundreds of publications in newspapers, magazines, radio, TV, web sites etc. around the world as well as in Slovenia have reported about these successes. Through our activities in the promotion of science we already have and will further contribute to inspire young people to study science and technology and try to increase public support for investments in knowledge. The research theme of the program is largely focused on health issues, where we are among the leading in the world in identifying mechanisms of defense against pathogens. We are transferring this knowledge into specific results that can in the future be used to improve human health, not only in Slovenia but also globally. Research in the field of synthetic biology is to a large extent an oriented basic project, where we always keep in mind the potential use of our studies in improving the quality of life, from improving the efficiency of biosynthetic production, reducing environmental burdens as well as energy demands of production to increasing production and improving efficiency, affordability and safety of new treatments. We are planning to convert the results obtained in our research into tangible high-tech products, which we intend to bring to the market in a variety of ways (starting companies, commercialising licenses and patents). Any potentially useful results will be protected by patents, which may allow us to sell licenses or start a business that would commercially exploit these results. Above all, we can expect a quick possibility for commercialization of new diagnostic methods and methods that improve a biosynthesis pathway.