Influence of salts on the preservation of microbial cell surface biosignatures exposed to UV radiation
Authors: Bourmancé Lucas , Ravaro Elisa , Toupet Maud , Nitsche Ruben , Brûle Sebastien , Raynal Bertrand , Elsaesser Andreas , Kish Adrienne 
Published: July 2024

High salt environments are ubiquitous in the solar system (Earth, Mars, Enceladus, Europa). As life on our planet is the only life we know of, terrestrial salt-loving (halophilic) microorganisms can be used to better characterize how life can thrive in such conditions. Halophilic archaea from the genus Halobacterium have been preserved in the fluid inclusions of halite crystals (NaCl) (Jaakkola et al., 2016). Hence, fluids inclusions may hold preserved biomolecules acting as ancient life biosignatures. Halites represent great exobiological interest as they have been identified on Mars (Osterloo, M. M. et al., 2008; Bramble &amp; Hand, 2024).Membrane lipids has already been described as good candidates for biosignatures because of their long-term stability properties (Georgiou &amp; Deamer, 2014) and membrane proteins, even though more fragile biomolecules, might be better preserved in high salt conditions.Therefore, this project aimed at understanding how Hbt. salinarum cell envelope fragments, produced by cell lysis, respond to UV irradiation (>185 nm) when incubated in different fluid inclusion compositions using a ground-based solar simulator. Fluid inclusions compositions were selected to represent Early Earth and Mars environments as well as modern Earth.High salt conditions are rarely compatible with traditional biochemistry methods and extensive optimization work is needed to render them suitable for the analysis of evaporite samples. For this project, this included optimization of cell envelope and membrane protein extractions, UV radiation exposure and investigating adequate methods for structural analysis.&#160;&#160;&#160;The chaotropic/kosmotropic effects of the brines on the structural stability of proteins and lipids of the cell envelope were determined using nano-Differential Scanning Fluorometry, Differential Scanning Calorimetry and Analytical Ultracentrifugation. In addition, a label-free mass spectrometry approach was employed to assess chemical modifications of membrane proteins (Orbitrap nano-LC-MS/MS) and lipids (GC-MS) after exposure to the different brines and radiation treatments. Finally, the photochemistry of the brines was investigated by looking at reactive oxygen species production using a fluorescent probe.This project has shown that certain brine ionic compositions are more prone to support cell envelope biosignature preservation against UV irradiation due to a combination of specific photochemistry and chaotropic effects. This work allows for the screening of various new methods for compatibility with high salts environment biochemistry required for analog studies on Earth but also for future analysis of space returned samples.The results of these ground-based experiments will be compared to real space irradiation as cell envelope samples will be exposed outside the International Space Station as part of the Exocube space experiment.This work was financed by the ANR ExocubeHALO ANR-21-CE49-0017-01 grant&#160;to A.Kish.&#160;References:Bramble, M. S., &amp; Hand, K. P. (2024). Spectral evidence for irradiated halite on Mars. Scientific Reports, 14(1). https://doi.org/10.1038/s41598-024-55979-6
Georgiou, C. D., &amp; Deamer, D. W. (2014). Lipids as universal biomarkers of extraterrestrial life. In Astrobiology (Vol. 14, Issue 6, pp. 541&#8211;549). Mary Ann Liebert Inc. https://doi.org/10.1089/ast.2013.1134
Jaakkola, S. T., Ravantti, J. J., Oksanen, H. M., &amp; Bamford, D. H. (2016). Buried Alive: Microbes from Ancient Halite. In Trends in Microbiology (Vol. 24, Issue 2, pp. 148&#8211;160). Elsevier Ltd. https://doi.org/10.1016/j.tim.2015.12.002
M. Osterloo et al. (2008), Chloride-Bearing Materials in the Southern Highlands of Mars, Science 319, 1651, DOI: 10.1126/science.1150690
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