The vision of RabXpress is the transfer of fundamental biological observations into applied projects with biotechnological potential. We employ a broad and state-of-the-art method portfolio including e.g. molecular genetics, synthetic microbiology and biochemistry.
In the past years the classical view of conventional eukaryotic secretion based on protein export via the endomembrane system has been challenged. The observation that an increasing number of important extracellular proteins is exported through alternative pathways has led to the discovery of fascinating and diverse export routes collectively termed ‘unconventional secretion’. In our group, we are interested in the unconventional secretion of the chitinase Cts1 in Ustilago maydis. In concert with the second but conventionally secreted chitinase Cts2, Cts1 mediates cell separation during cytokinesis. A lack of the two proteins results prevents cell separation and thus leads to cellular aggregation. While the function of Cts1 has been demonstrated by our group, its recruitment to the fragmentation zone connecting mother and daughter cell remains unclear. Recently, we identified Jps1, a protein that plays an important role for Cts1 secretion and accordingly cell separation. We currently aim at deciphering the molecular details of this unconventional secretion mechanism by tackling the individual proteins and their crosstalk.
This project is associated with the CRC1208.
Exploiting unconventional secretion for pharma protein export
Protein secretion is highly attractive for the production of recombinant proteins with biotechnological application as it significantly reduces downstream processing costs. However, proteins travelling via the endomembrane system are frequently modified by post-translational modifications like N-glycosylation which can be disruptive for their activity in heterologous hosts. To avoid these modifications, we employ unconventional secretion in U. maydis for the export of hydrolytic and pharmaceutical proteins like nanobodies, single-chain variable fragments or carbohydrate-active enzymes (CAZymes). Here, we employ the aforementioned Cts1 as a carrier. A unique feature of the system is that Cts1 binds to chitin which can be employed for purification. To establish the fungus as an expression host, we are currently tuning the yields of the system and optimize the system at different levels. In parallel we evaluate the option of surface immobilization via chitin interaction. Potential future applications of our platform are versatile virus detection systems based on inexpensive chitin surfaces or production of antibody formats for pharma.
As a plant pathogen U. maydis harbours a small but potent set of hydrolytic enzymes for the degradation of biomass polymers. In the past years we successfully enhanced the capacity of the fungus for degradation of biomass components like cellulose or polygalacturonic acid. We used a combination of genetic activation of intrinsic enzymes and complementation with potent foreign enzymes via conventional and unconventional secretion. In addition, U. maydis is a natural producer of valuable molecules including glycolipids, polyols, organic acids and hydrolytic enzymes. We also engineered the fungus for production of foreign molecules like sesquiterpenes and pharmaceutical proteins via unconventional secretion. In the future we will combine both properties and produced valuable compounds from plant biomass waste and industrial side- and waste streams.
Several applied projects are associated with the BioSC.
Synthetic communities mimicking lichen
In nature microbes are usually embedded in intimate interaction networks with other microorganisms. One prime example is the lichen symbiosis, a very ancient and tight association of algae and/or cyanobacteria with asco- and basidiomycete fungi. The current understanding of such a cross-kingdom community is poor. We aim to get deeper insights into the underlying molecular principles by employing synthetic communities of cyanobacteria and fungi. By targeted manipulation using synthetic biological tools we will modulate the interaction at the level of carbon metabolism and physical interaction and read-out the physiological consequences. Using this principle, we will gather information on the organisation of organismic interaction networks in order to assemble new associations from scratch in the future.
Exploring synthetic communities for biotechnological applications
Microbial communities have major advantages in comparison to pure cultures. For example, they follow the strategy of work sharing and complementary metabolism. This decreases the individual burden of the cells. In a biotechnological approach we aim to exploit lichen-like communities for the production of valuable molecules from sunlight. Here, engineered phototrophic cyanobacteria grow on light and CO2 and produce sucrose to feed heterotrophic fungi like U. maydis or S. cerevisiae. Depending on the associated fungi, different valuable substances will be produced from sunlight.
M. Philipp, K. P. Hussnaetter, M. Reindl, K. Müntjes, M. Feldbrügge and K. Schipper (2022) A novel potent carrier for unconventional protein export in Ustilago maydis. Front. Cell Dev. Biol. 9: 3825
N. Wierckx, K. Miebach, N. Ihling, K. P. Hussnaetter, J. Büchs and K. Schipper (2021) Perspectives for the application of Ustilaginaceae as biotech cell factories. Essays Biochem. 65: 365–379
P. Stoffels*, M. J. Müller*, S. Stachurski, M. Terfrüchte, S. Schröder, N. Ihling, N. Wierckx, M. Feldbrügge , K. Schipper# and J. Büchs# (2020). Complementing the intrinsic repertoire of Ustilago maydis for degradation of the pectin backbone polygalacturonic acid. J. Biotechnol. 10: 148-163
* shared first authorship # shared corresponding authorship
M. Reindl*, J. Stock*, K. P. Hußnätter, A. Genc, A. Brachmann, and K. Schipper (2020) A novel factor essential for unconventional secretion of chitinase Cts1. Front. Microbiol. 11: 1529
*shared first authorship
Research group leader