Team:Concordia-Montreal/Poster

Poster: Concordia-Montreal



Title

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AstroBio, open-source database for gene expression in microgravity and AstroYeast, resistant strains to microgravity-induced stress

Presented by Team Concordia Montreal 2020

Team : Lancia Lefebvre, Hajar El Mouddene, Paula Gomez, Maher Hassanain, Gabriel Aguiar-Tawil, Evelyn Huaman, Grecia Olano O'Brien, Samman Zaman, Benjamin Clark, Labrini Vlassopoulos, Natasha Letourneau, Brian Baxter, Amin Nikpayam, Khashayar Zardoui, Nhi Hoang Nguyen, Asif Iqbal.

Abstract

Advances in synthetic biology, including cellular agriculture, enable the sustainable production of food on Earth. However, in-space biomanufacturing, for which maintaining cultures in bioreactors for extended periods is essential, has proved challenging. Microgravity induces global changes in gene expression profiles, triggering stress responses in cells. For example, Saccharomyces cerevisiae exhibits stress responses characterized by aberrant cell polarity, budding, and separation, which affects cell growth and productivity in space. There is also a lack of bioinformatics tools for microgravity researchers. To fill this gap, we developed AstroBio, an open-source database compiling literature findings on microgravity-induced gene expression changes in different model organisms. The database informs our development of AstroYeast, yeast strains that are resistant to microgravity-induced stress. This will be done in a high-throughput manner either by strain adaptive evolution, or genome-wide overexpression and knockdown screens. AstroYeasts can be used to sustainably and renewably produce nutrients in space under microgravity conditions.

AstroBio & AstroYeast a Two-Year Project
We wanted to find a solution using synthetic biology and we thought of AstroYeast which are space-compatible strains for biomanufacturing of nutrients in space. Meaning they are optimized for bioproduction in space, as they are resistant to microgravity-induced changes in gene expression. For our research, we needed a database that would allow us to perform robust analyses of current literature findings but we noticed a lack of bioinformatic tools for microgravity research. As a result, we decided to build a database to fill this gap. We then used our AstroBio database to directly inform the development of AstroYeast, which will be delivered next year.
Inspiration

Last May, our team witnessed an awe-inspiring moment for humankind when the first crewed SpaceX flight launched to the International Space Station (ISS). This historic launch is part of a larger effort to commercialize Low Earth Orbit as space agencies have now shifted their agendas to exploring the next frontiers in the universe.

In 2024, NASA and the ISS partners, including Canada, are planning to launch the lunar gateway, a space station in lunar orbit intended to serve as a communication hub, science laboratory, a short-term habitation module, and a critical gateway to expanding a human presence on Mars.

SpaceX is also planning to launch the first crewed flight to Mars in 2024. At iGEM Concordia, we are inspired by the discoveries humankind will make on these journeys as our scientists conduct research in altered gravity conditions.

To perform research in space and explore the universe, we need to provide sustenance to the crews. On a space station, the environment is limited in terms of available space and resources such as water and oxygen.


How will astronauts’ nutritional needs be fulfilled on multi-year round trips to Mars? Or for long- term sustainable habitation on space stations?

This is what inspired AstroYeast: space-compatible strains for biomanufacturing applications in space. Our database, AstroBio, was a direct product of our background research to build for AstroYeast. We needed a web application that would enable us to directly compare experimental findings on microgravity-induced gene expression changes in yeast and to determine whether these changes are microgravity-specific when compared to other stressors. However, we could not find any bioinformatics tools that would enable us to perform the analysis that we needed. As a result, we decided to create our software and database, AstroBio.

Problem

In-space biomanufacturing, for which maintaining cultures in bioreactors for extended periods is essential, has proved challenging. Microgravity induces global changes in gene expression profiles, triggering a stress response in cells. For example, Saccharomyces cerevisiae exhibits a stress response characterized by aberrant cell polarity, budding, and separation which affects cell growth and productivity in space. To develop microgravity-tolerant, space-compatible yeast strains as a chassis for biomanufacturing in space, we needed to select a strong set of gene promoter candidates whose expression is significantly, reliably and specifically affected by microgravity-induced stress. The problem is that at the moment:

  • There exists a large and increasing amount of unorganized data on how living organisms react to environmental conditions in space. There is a need for a well-curated, microgravity-specific database to analyze and organize the increasing amount of microgravity-related gene expression data
  • There is no database specific to microgravity research. Available databases are restricted to specific types of experiments including analysis of either RNA-seq or microarray datasets
  • Databases are limited in terms of their search capabilities and they do not allow researchers to directly and easily compare experimental findings across studies.
Idea
We wanted to find a solution using synthetic biology and we thought of AstroYeast which are space-compatible strains for biomanufacturing of nutrients in space. Meaning they are optimized for bioproduction in space, as they are resistant to microgravity-induced changes in gene expression. For our research, we needed a database that would allow us to perform robust analyses of current literature findings but we noticed a lack of bioinformatic tools for microgravity research. As a result, we decided to build a database to fill this gap. We then used our AstroBio database to directly inform the development of AstroYeast, which will be delivered next year.
Human Practices
At iGEM Concordia, we understand that while the creation of these solutions holds a promise for a better future, it also holds a responsibility to ensure that these innovations are valid, safe, ethical, and representative of stakeholder needs and concerns. Throughout the development of AstroBio and AstroYeast, we consulted with a number of academic researchers, industry experts, space agencies, and end-users to validate the problem that we have identified and to ensure that our solution is not only of value to these stakeholders but that it is also ethical and safe for our community. Stakeholder needs and concerns were documented and the feedback that we received was used to inform the design and execution of our project. Our approach to human practices is summarized in the following three-step process:

  1. Connect:We connected with academic researchers, space agencies, space and biomanufacturing industry, and end-users.
  2. Exchange:We conducted an hour-long Zoom interview with each one of our stakeholders and documented our exchange.
  3. Integrate:We discussed the feedback that we received and implemented changes to the development of AstroBio and AstroYeast.

User-Centered Design
To ensure engineering success we consulted directly with end users and gathered feedback from AstroBio testing.

We had Macauley Green from the Astropharmacy & Astromedicine department at the University of Nottingham test our AstroBio database. He is interested in well-curated data, data visualisation, knowing the change in expression in microgravity conditions, and inclusion generation information. Macauley suggested a search criteria which would distinguish between the study type, such as a study in simulated microgravity here on Earth versus a space flown experiment that was performed on the International Space Station.

We believe that integrating our Human Practices makes for solutions which we would have not come to on our own in such a short time frame.
AstroBio Saccharomyces cerevisiae MultiStress Explorer
We needed to narrow down the initial gene selection to those that are specifically regulated by microgravity-induced stress, rather than by other stressors such as heat shock. We were unsure how to do this and sought out expert advice from bioinformaticians Mathieu Harb and Richard Barker. They suggested using heat maps to visualize the data and provided advice on statistical and visualization methods and resources that helped us build the AstroBio MultiStress Explorer.

It is a small web-application for visualizing transcriptomics data from Saccharomyces cerevisiae in multiple stress conditions, including microgravity-induced stress.

Researchers can search a specific gene or multiple genes, choose between stressors comparisons: heat shock, high osmolarity, and oxidative stress. Results can be visualized in heatmaps, volcano plots, gene forest plots, and PCA-biplots. We used AstroBio and the MultiStress Explorer to find yeast genes that are significantly, specifically and reliably upregulated in microgravity. The genes we selected informed the design of our space-compatible AstroYeast strains.
AstroBio, an open-source database for gene expression in microgravity
well-curated, open-source, and user friendly software and database compiling literature findings on microgravity-induced gene expression changes in yeast, bacteria, and plants.

Astrobio features

  • Specialized microgravity database
  • Well-curated data
  • No data-type restrictions
  • User-friendly
  • Allows direct comparison of findings across studies
  • Open source

We continuously integrated our user’s needs throughout our project design.

We included Essential Genes for yeast survival in microgravity thanks to Dr. Nislow.

We performed exploratory analysis and incorporated visualisations of our data as requested by many microgravity researchers and bioinformaticians. Specifically we included heat maps, PCA biplots, volcano and gene forest plots and developed the MultiStress Explorer.

From iGEM Toulouse’s feedback we improved the user interface of AstroBio as we added sorting, pagination, and limitations to the number of results displayed. We also changed the results to display to a more user-friendly view. From Macauley Green’s feedback, we validated our choice to include generation as a factor in our gene expression analysis and performed an exploratory analysis to determine if there are generation effects across experimental findings.

We validated that other researchers such as Morgan Irons, who launched an experiment to the ISS this September, would use and contribute experimental findings to AstroBio.

Next year, we will continue to collaborate with Dr. Richard Barker and we will improve AstroBio as we:
  • Add more studies and include more organisms
  • Explore pathway visualization
  • Upset R plot to compare and explore the overlap between studies
  • Allow researchers to submit their published data to our database
From AstroBio to AstroYeast space-compatible strains
Our goal is to create Saccharomyces cerevisiae strains that are tolerant to microgravity-induced stress for bio manufacturing applications in space. To achieve this we used the AstroBio database to select promoters whose expression is highly and consistently upregulated in microgravity. These will be used as the promoter for a GFP-based reporter system that will be used to track the changes in expression of these genes due to microgravity stress. The reporter will be constructed within the yeast using homologous recombination and inserted using the CRISPR-Cas9 system. We will use a microgravity simulator as a way to apply the microgravity stress to the yeast. This will be used as the stressor for adaptive evolution experiments that will lead to the eventual resistance to microgravity. As a proof of concept, the genes necessary for vitamin A production will be inserted into the resistant yeast to see if it has an effect on the production of this biomolecule.

Promoter Selection

  1. We selected promoters with upregulated expression in microgravity
  2. We refined our selection criteria to promoters with expression upregulation that is specific to microgravity
  3. We further refined our selection criteria to promoters in pathways which have been shown to be significantly affected by microgravity
  4. We added a promoter for Vitamin A.In partnership with iGEM Toulouse, we included pGAL10 as a proof of concept for Vitamin A production in yeast under microgravity conditions.
References & Attributions
We are excited to continue the development of an R&D platform to facilitate experimental research and biomanufacturing applications in space. Our team comes from a diverse background which is reflected in the breadth of work we have done this year. Our team gives light-years of thanks to everyone who has contributed to the realisation of AstroBio and AstroYeast including our instructors, mentors, and sponsors.

Visit our wiki for more details: by clicking here

References
  1. Afgan, E., D. Baker, B. Batut, M. vandenBeek, D. Bouvier, M. Čech, J. Chilton, D. Clements, N. Coraor, B.A. Grüning, A. Guerler, J. Hillman-Jackson, S. Hiltemann, V. Jalili, H. Rasche, N. Soranzo, J. Goecks, J. Taylor, A. Nekrutenko, and D. Blankenberg. (2018). The Galaxy platform for accessible, reproducible and collaborative biomedical analyses: 2018 update. Nucleic Acids Res.46:W537–W544. doi:10.1093/nar/gky379.
  2. Buckley, N., & Johnson-Green, P. (n.d.). Microgravity Research in the Canadian Space Agency. Acta Astronautica, Volume 63, p. 35-37.
  3. Davis, S., and P.S. Meltzer. (2007). GEOquery: a bridge between the Gene Expression Omnibus (GEO) and BioConductor. Bioinformatics. 23:1846–1847. do.i:10.1093/bioinformatics/btm254
  4. Menezes, A., Cumbers, J., Hogan, J., & Arkin, A. (2015). Towards synthetic biological approaches to resource utilization on space missions. Journal of the Royal Society Interface.
  5. MongoDB inc. (2020). Choose which type of deployment is best for you. Retrieved from MongoDB
  6. Nislow, C., A.Y. Lee, P.L. Allen, G. Giaever, A. Smith, M. Gebbia, L.S. Stodieck, J.S. Hammond, H.H. Birdsall, and T.G. Hammond. (2015). Genes Required for Survival in Microgravity Revealed by Genome-Wide Yeast Deletion Collections Cultured during Spaceflight. BioMed Research International. 2015:e976458. doi:10.1155/2015/976458
  7. Sheehan, K. B., McInnerney, K., Purevdorj-Gage, B., Altenburg, S. D., & Hyman, L. E. (2007). Yeast genomic expression patterns in response to low-shear modeled microgravity. BMC genomics, 8, (3). Doi: 10.1186/1471-2164-8-3


Poster designed by Paula Gomez, Maher Hassanain, Gabriel Aguiar-Tawil and Lancia Lefebvre. With contributions to visuals and content from Hajar El Mouddene and Grecia Olano O’Brien.