Why is our project worth the award for the best plant synthetic biology project?
Mossphate provides first insights in the biotechnological approach to close the phosphate cycle by using the moss Physcomitrella patens (P. patens). The overuse of fertilizers on crop plant fields causes the phosphate and other nutrients to sink into the groundwater and therefore reach lakes and rivers, where mainly the saturation of phosphate and nitrogen causes eutrophication (Prasad & Prasad, 2019). This leads to numerous problems. Fertilizers are produced by exploiting phosphate rocks, which decrease over the years because of the increasing demand in fertilizers (Cordell, D., Tina schmid-Neset, Stuart White, 2009; Cordell, 2010; Cordell et al., 2009).
P. patens is a promising candidate for our approach, as it shows great potential in possible applications. To begin with, it is a model organism currently used in different fields of research regarding plant biology. Its photoautotrophic metabolism represents a clear advantage since it does not require carbon in its medium to be able to grow. In addition, it can be cultivated in both solid medium and liquid culture. A key characteristic which makes P. patens a perfectly apt candidate for Mossphate is it’s high degree of homologous recombination, which enables a precise editing of genomic DNA (Frank, Decker, and Reski 2005).
The final goal of our project is to engineer this moss for an enhanced uptake of phosphate out of wastewaters and its accumulation in the form of polyphosphate granules. With this, we aim to prevent eutrophication of lakes and rivers. The moss, which will be rich in polyphosphate, would ideally be used as fertilizer.
Improve phosphate storage in the form of polyphosphate granules.This can be achieved by overexpressing the native polyphosphate kinases (PPKs) or by heterologous expression of PPK variants from E. coli. Those PPKs convert the excess inorganic phosphate (Pi) into osmotically inert polyphosphate granules (Rudat, Pokhrel, Green, Gray 2018).
Test the accumulation of polyphosphate in the cytosol and vacuole.In some cases the high accumulation of polyphosphate leads to toxicity problems for several organisms (Rudat, Pokhrel, Green, Gray 2018). To better assess the effect on toxicity in our P. patens system, the localization of polyphosphate granules in the cytosol and vacuole could be tested by expressing the PPKs with and without the presence of a vacuolar import signal.
Enhance phosphate uptake capacity of P. patens.The uptake ability of P. patens could be tested by the overexpression of native phosphate transporters, more concretely PpPHO1;1 and PpPHO1;7 (Wang, Secco, & Poirier, 2008).
Due to the covid-19 Pandemic, labs at our university were shut down and we had only minimal time available for a proof of concept. We cultured the moss, but were not able to genetically modify it in time for further experiments. We did however, lay the groundwork for metabolic modelling and genetic engineering in P. patens, an underrepresented chassis in iGEM and plant biology in general.
P. patens will be integrated into the wastewater facilities (WWF) by integrating a net step consisting of a turf-scrubber photobioreactor. There, the moss will be grown on a perforated surface, anchoring itself by rhizoids. This way, the mosses in the reactor can be easily exchanged for fresh ones by simply exchanging the surface. The safety of the system can be ensured by the integration of genetic kill-switches such as red-light specific proteases used by Team Munich 2013. Those proteases recombine when being illuminated to cleave a membrane-bound nuclease, which later translocates into the nucleus and degrades the genome (iGEM Munich, 2013). The moss would normally be protected by a filter, however when escaping to the environment would no longer be protected resulting in the self-destruction of the moss.
Cordell, D. (2010). The Story of Phosphorus Sustainability implications of global phosphorus scarcity for food security. Environmental Studies. Retrieved from http://swepub.kb.se/bib/swepub:oai:DiVA.org:liu-53430?tab2=abs&language=en
Cordell, D., Drangert, J.-O., & White, S. (2009). The story of phosphorus: Global food security and food for thought. Global Environmental Change, 19(2), 292–305. https://doi.org/10.1016/J.GLOENVCHA.2008.10.009
Frank, W., Decker, E. L., & Reski, R. (2005). Molecular tools to study Physcomitrella patens. Plant Biology, 7(3), 220–227. https://doi.org/10.1055/s-2005-865645
Prasad, R., & Prasad, S. (2019). Algal Blooms and Phosphate Eutrophication of Inland Water Ecosystems with Special Reference to India. International Journal of Plant and Environment, 5(01), 1–8. https://doi.org/10.18811/ijpen.v5i01.1.
Rudat, A. K., Pokhrel, A., Green, T. J., & Gray, M. J. (2018). Mutations in Escherichia coli polyphosphate kinase that lead to dramatically increased in vivo polyphosphate levels. Journal of Bacteriology, 200(6). https://doi.org/10.1128/JB.00697-17
Wang, Y., Secco, D., & Poirier, Y. (2008). Characterization of the PHO1 gene family and the responses to phosphate deficiency of Physcomitrella patens. Plant Physiology, 146(2), 646–656. https://doi.org/10.1104/pp.107.108548