Team:Sorbonne U Paris/Description

THE “BIOILOGICAL FACTORY” PROJECT

Our 2019 iGEM team is composed of students from Sorbonne Université (Paris, FRANCE), initially drawn from diverse backgrounds but sharing a common passion for Science and Innovation. Our project entitled the “Bi[oil]ogical Factory”, consists in developing a new chassis for the production of lipid compounds, in a photosynthetic green microalga named Chlamydomonas reinhardtii. We want to build our proof-of-concept by modifying this microorganism to produce palmitic acid (C16:0) and oleic acid (C18:1), which are the main components of palm oil.

HOW ?

Given that palmitic acid and oleic acid are the products of the same metabolic pathway, we decided to develop two different strains of C. reinhardtii, each one producing one of the two fatty acids of interest. On one hand, we will express FAT-B2, LPAAT-A and DGAT1-2 from the palm oil Elaeis guineensis in C. reinhardtii using the Golden Gate Modular Cloning (MoClo) technology, based on Type IIS restriction enzymes1 to enhance the proportion of triacylglycerols (TAG) containing palmitic acid (C16:0). The first enzyme catalyzes the release of the free fatty acid from the acyl-coA and the second and third enzyme catalyzes the incorporation of the second and third fatty acid on the glycerol to produce TAGs. We have chosen these enzymes because they potentially determine the fatty acid composition of TAG in the mesocarp, the pulp of the palm fruit2. On other hand, we will express FAT-A, LPAAT-A and DGAT1-2 from E. guineesis in C.reinhardtii to enhance the proportion of TAG containing oleic acid (C18:1). In addition, we will knock-down the enzymes (fatty acid desaturases) that use oleic acid as a substrate for the production of linoleic acid (C18:2) and linolenic acid (C18:3): allowing the accumulation of oleic acid (C18:1). This step will be done by using the RNA interference technology. Finally, to enhance TAG production we will grow Chlamydomonas reinhardtii in nitrogen starvation conditions.

WHAT INSPIRED US ?

We were initially drawn to this project by the environmental issues caused by the massive palm oil production. Palm oil is extensively used by many industries such as the food industry, the biofuels and even in cosmetics. The main producing countries such as Indonesia and Malaysia have seen an increase of the pollution caused by the progressive and massive deforestation. Malaysia has lost approximately 48 000 km² of its forest between 2000 and 20123, so the biodiversity has been extremely disturbed . For instance, it leads to the destruction of habitats of certain species like orangutan. Moreover, other ethical issues are at stake such as the lack of respect for human rights since the production involves the exploitation of young children according to Amnesty International. Therefore, we were investigating a new model that could be modified by synthetic biology tools to produce the same lipid compounds as those found in palm oil which would not require an oil extraction from palm tree, in other words would not be a threat to the environment. We decided to use Chlamydomonas reinhardtii as a chassis for this production because it is a mixotroph (i.e. both autotrophic and heterotrophic) and it has the ability to accumulate TAG in the form of lipid droplets and the capacity to produce less carbon than it consumes. Moreover, its culture does not imply the occupation of arable lands. So, we decided to extend our project to the development of a new chassis, Chlamydomonas reinhardtii for the production of lipid compounds. Moreover, interviews with experts helped us in the building of our strategy to enhance the production of our lipid compounds of interest: palmitic acid and oleic acid. After an interview with Jean-Luc Cacas, (a french researcher at the INRA - National Institute of Agronomical Research) we decided to take our inspiration from a strategy that has successfully worked in plants, the “Push, Pull and Protect” strategy4. It involves optimizing the flux of carbon into TAG (storage form of lipids) by increasing the fatty acid synthesis (Push), TAG assembly (Pull) and lipid turnover (Protect).






Followed by the interview with Juliette Puyaubert, (a french lecturer specialized in the lipid metabolism of plants) that led us to focus on three important enzymes. The first one is FAT (fatty acid thioesterase) which plays a role in the export of fatty acids from the chloroplast to the endoplasmic reticulum5. The second one is LPAAT (lysophosphatidic acid acyltransferase) that is important for fatty acids assembly into TAG5. The last one is DGAT (diacylglycerol acyltransferase), it seems to be overexpressed in the mesocarp of palm oil6.






In addition to this, from the designing step to the achievement of our project we aim to spread scientific knowledge and innovation in the field of biology, to both scientists and nonscientists: through articles written in a popular science journal, the designing of a video game and a comic book. 
Moreover, we have planned as many cultural events like scientific activities around synthetic biology and the Bi[oil]ogical Factory project.



For more details, go on our Education and Public Engagement section :


EDUCATION & PUBLIC ENGAGEMENT




  1. Crozet, P. et al. Birth of a Photosynthetic Chassis: A MoClo Toolkit Enabling Synthetic Biology in the Microalga Chlamydomonas reinhardtii. ACS Synth. Biol. 7, 2074–2086 (2018).
  2. Dussert, S. et al. Comparative transcriptome analysis of three oil palm fruit and seed tissues that differ in oil content and fatty acid composition. Plant Physiol. 162, 1337–1358 (2013).
  3. M. C. Hansen et al., « High-Resolution Global Maps of 21st-Century Forest Cover Change », Science (New York, N.Y.), vol. 342, p. 850‑853, nov. 2013.
  4. Vanhercke, T. et al. Metabolic engineering of biomass for high energy density: oilseed-like triacylglycerol yields from plant leaves. Plant Biotechnol. J. 12, 231–239 (2014).
  5. Li-Beisson, Y., Beisson, F. & Riekhof, W. Metabolism of acyl-lipids in Chlamydomonas reinhardtii. The Plant Journal 82, 504–522 (2015).
  6. Xiao, Y., Xia, W., Mason, AS., Cao, Z., Fan, H., Zhang, B., Zhang, J., Ma, Z., Peng, M., Huang, D. Genetic control of fatty acid composition in coconut (Cocos nucifera), African oil palm (Elaeis guineensis), and date palm (Phoenix dactylifera). Planta. 2019 Feb, 249 (2), 333-350