iGEM 2016: EPFL team wins gold medal, receives two honorable mentions

The International Genetically Engineered Machine (iGEM) competition takes place each year, with teams of high-school and university under- and post-graduate students compete with synthetic biology projects. It is organized by the iGEM foundation, an independent non-profit “dedicated to education, competition, the advancement of synthetic biology, and the development of an open community and collaboration.” For most teams, the competition runs for the larger part of the year culminating at the iGEM Giant Jamboree in Boston, MA.

The 2016 team from EPFL, composed of nine 3rd year SV Bachelor students, was led by Bart Deplancke, Barbara Grisoni, and further aided by two PhD students of the Deplancke Lab: Roel Bevers and Riccardo Dainese.

The students were:

• Alix Faillétaz
• Dimitri Coukos
• Camilla Ceroni
• Marie Matos
• Yassine Zouaghi
• Rémy Pétremand
• Francesco Terenzi
• Pietro Airaghi
• Samuele Mercan

The project that the team worked on is dubbed “Intelligene” and uses as principal component dCas9, which is an inactivated or “dead” version of the Cas9 endonuclease widely used in CRISPR gene editing techniques. As an inactive mutant, dCas9 can no longer edit DNA but can still bind to double-stranded DNA at virtually any locus through a guide RNA (gRNAs). As such, dCas9 can be used as a “programmable” transcription factor, which is the central idea behind Intelligene. For example, gRNAs can be designed that guide dCas9 to activating regions (promoter) of a gene. Once bound, transcriptional activators or repressors can be recruited so that dCas9, together with these co-factors, acts as an “on” or “off” switch of specific genes in a “decision-making” cell circuit.

“We built upon efforts of the 2015 team’s work,” says Alix Faillétaz. “They found that the building of such a decision-making circuit wasn’t really reliable in yeast using the classical dCas9 system, so we decided to try a new, much more modular system.”

The notion of using dCas9 as a “programmable” transcription factor is not new; what is, however, is using it to build intuitive and complex transcriptional networks in cells, which can be programmed and modified to perform specific biological functions.

“The previous system (with the transcriptional effector fused to the dCas9) wasn’t the best tool for biological circuit creation as you always have the activation unit fused to the dCas9, even when you want to repress a gene”, says Alix Faillétaz. “So we further characterized in yeast a system developed by Zalatan et al. in 2015 (“Engineering Complex Synthetic Transcriptional Programs with CRISPR RNA Scaffolds”), who added a hairpin to the gRNA to recruit either an activator or a repressor, depending on the effect you want on the region targeted by the gRNA. As a proof of concept, we built a functional and galactose-inducible NOT gate.”

The new dCas9 scaffold system used by the 2016 iGEM team. The gRNA is supplemented with an additional hairpin (scaffold) so that it can recruit transcriptional effectors. This effectively allows it to become a scaffold RNA (scRNA). In this way, the scRNA does not only determine the sequence of the promoter targeted, but also the effect imposed on the promoter.

The team didn’t just stop at biology; there was a software part to the project as well. “We worked off a program developed at MIT that helps build circuits in any system using a network of logic NOR gates,” says Dimitri Coukos. “The program shows you the best option, but it only works with actual transcription factors and not synthetic ones such as dCas9-based version.” This last aspect was one of the challenges the team wanted to solve: transcription factors are too site-specific and tend to interfere with a cell’s own genetic network.

“Our system does not interfere with the normal activity of the cell compared to systems based on actual transcription factors,” explains Samuele Mercan. “We’re not editing the genes, we’re just inserting a new regulatory code.”

“The bio part of the project allows you to create less toxic circuits and design them more easily,” continues Coukos. “That’s why it’s called Intelligene — it’s an intelligent design of gene circuits.”

“The aim, from the beginning, was to design both the biological and software components in a way that facilitates the use of the system for the scientific community,” says Marie Matos. For that purpose, the team made sure that the software is open-source, and accessible online by other labs.

“The Intelligene system is a framework for designing behavior in cells,” says Dimitri Coukos. “We took the software and made it more compatible with our system. One of the problems was that the MIT version uses a very specific code for user input, which we replaced with a drag-and-drop graphical user interface. Then we modified the file structure and moved it from a local system to the cloud, to enable scientists around the world to collaborate.”

The final Jamboree ran from 27-31 October in Boston’s Hynes Convention Center, hosting 5,600 participants making up 300 teams from 42 countries (full results). Though the Grand Prizes went to Imperial College (Undergrad) and LMU-TUM Munich (Overgrad), the EPFL team was awarded a gold medal for their work and nominated in two categories: best information processing project and best software tools.

“The iGEM competition was one the most tiring but amazing experiences ever,” says Rémy Pétremand. “One of the judges critiqued our work but he also liked it very muchwas also very enthusiastic about it and gave us great comments about how we could improve it.”

“We met a lot of teams, networked, and shared our experiences,” says Camilla Ceroni. “We even interviewed a lot of teams and it was cool to talk to them and also learn from their work,” adds Yassine Zouaghi.

“A more immediate application would be to develop biosensors of programmed (yeast) cells to detect toxins in water or heavy metals in the soil,” says Bart Deplancke. “But of course we also want to go beyond yeast and into mammalian cells.” And there might also be a paper in the works: “We’ve never been this close with an iGEM team to publishing,” he adds, to sheepish grins and “we’ll see” shrugs from the team, “but what the students accumulated in terms of research experience and project management is already priceless”.