NOTE: End date changed to 04/29/2024 per NSSC information (Ed., 5/31/23)

NOTE: End date changed to 04/30/2023 per NSSC information (Ed., 4/12/23)

The first aim of this project is to identify the metabolic and genomic pathways in yeast affected by microgravity, space radiation, and a combination of both. The second one is to differentiate between gravity and radiation exposure on single-gene deletion mutants’ ability to thrive in the spaceflight environment. We hypothesize that mutants lacking genes associated with DNA repair, recombination, and replication will have lower survivability rates beyond the van Allen belts than their below van Allen belts- or Earth-controls

The experiment is designed to have a controlled start after Orion is past the van Allen belts, grow ~21 generations of the deletion series, and fix or preserve samples for post-flight analyses. Should the automated controlled approach be considered inappropriate for implementation on EM-1, we have a passive approach that is based on dotting each mutant individually on agar. We have performed both approaches in space in the past.

This project will address three Space Biology Program Science Elements, three Objectives, three Guiding Questions, and four Decadal Survey’s highest priority Recommendations by preserving nucleic acids of different generations of the yeast deletion series cultures grown in space, beyond as well as below the van Allen belts (and uploading the genomic and transcriptomic data to GeneLab).

Books

1. Pathak, Y., Araújo dos Santos, M., Zea, L. Handbook of Space Pharmaceuticals. Springer International Publishing

Peer-Reviewed Journal Papers

2. Gaikani, et al., One Small Step for Yeast — NASA’s Artemis I mission reveals Lunar Spaceflight induces DNA methylation regulating transcription in yeast, under review, Cell 3. Zea, L., Piper, S. S., Gaikani, H., Khoshnoodi, M., Niederwieser, T., Hoehn, A., ... & Nislow, C. (2022). Experiment verification test of the Artemis I ‘Deep Space Radiation Genomics’ experiment. Acta Astronautica, 198, 702-706. https://doi.org/10.1016/j.actaastro.2022.06.018 Conference Papers and Posters, Open-Access Code, Podcasts, and STEM outreach 4. Zea, L., Levine, H., Santamaria, S., Hammond, T., Brandizzi, F., Henessa, T., Artemis I Biological Payloads Panel, ASGSR 2023 5. Zea, L., Nislow, C., Artemis I’s Deep Space Radiation Genomics Opportunities, Constraints, & Challenge Solving, NASA Artificial Intelligence / Machine Learning (AI/ML) Working Group, April 10, 2023 6. Zea, L., Niederwieser, T., Hoehn, A., Grusin, M., Wright, J., Stanish, G., Piper, S., Flores, P., Griffith, R., Gaikani, H., Khoshnoodi, M., Siems, K., Cortesao, M., Wahl, K,. Bernstein, V., Knipp, D., Wilson, K., Scott, K., Lutsic, A., Soler, M., Sato, K., Viau, A., Aragon, R., Stodieck. L., Countryman, S., Carr, C.E., Moeller, R., Nislow, C., From Idea to Lunar Orbit Flight Readiness - Implementation of the Artemis I 'Deep Space Radiation Genomics' (DSRG) Yeast Experiment, American Society for Gravitational and Space Research (ASGSR) Conference, November 3-7, 2021 7. Zea, L., Piper, S., Gaikani, H., Khoshnoodi, M., Niederwieser, T., Hoehn, A,. Grusin, M., Wright, J., Flores, P., Wilson, K., Lutsic, A., Stodieck, L., Carr, C.E., Moeller, R., Nislow, C., Experiment Verification Test of the Artemis I 'Deep Space Radiation Genomics' Experiment, IAC-21-A.2.7x65012, 72th International Astronautical Congress, Dubai, United Arab Emirates 8. Zea, L., Nislow, C., First Controlled Biological Experiments beyond the Van Allen Belts, American Institute of Aeronautics and Astronautics (AIAA) Rocky Mountain Chapter Annual Technical Symposium, University of Colorado Boulder, September 29, 2021 9. Bernstein, V. and Wahl, K. (2021). Identifying and Visualizing the Moon's Position with respect to the Earth's Magnetotail (v1.0.0). Zenodo. [Github] 10. Wahl, K.R., Bernstein, V., Knipp, D., and Zea, L., Maximizing Space Radiation Exposure for the Deep Space Radiation Genomics (DSRG) Experiment on Artemis 1, 18th Space Weather Conference in the American Meteorological Society 101st Meeting 10-14 January 2021 (Virtual) 11. Fiske Planetarium Podcast “A View From Earth”, Episode 8 - Life finds a Way - Or does it? Guests: Luis Zea & Bruce Jakosky, July 2020 [Podcast] [Video] 12. Wahl, K., Bernstein, V., Knipp, D. Deep Space Radiation Genomics (DSRG) Experiment on Artemis 1: Magnetotail and Bow shock Affect, Research Experience for Undergraduates (REU) Final Presentations, University of Colorado, Boulder, July 29, 2020 13. Meda, E., Herbruger, E., Hernandez, D., Aragón, R., Aguirre, D., Alvarado, I., Zea, L., Viau, A., Piezas para proyecto DSRG - Artemis 1, UVG 2020 Science Fair 14. Knipp, D., Wahl, K., Bernstein, V., Zea, L. Sending Tiny Life Forms to Space. ED032 - The Up-Goer Five Challenge: Explaining big important things in simple words. AGU 2020 Fall Meeting, December 1-7, 2020 15. Breaux S, Fuchs F, Cortesao M, Siems K, Stodieck L, Niederwieser T, Zea L, Carr CE, Moeller R, Nislow C. "Irradiation Ground Control for a Genome-Wide Yeast Fitness Profiling Experiment On Board Orion’s Artemis 1 Mission." 35th Annual Meeting of the American Society for Gravitational and Space Research, Denver, CO, November 20-23, 2019. 16. Zea L, Niederwieser T, Stodieck L, Carr C, Moeller R, Nislow C. "Experiment Design for a Genome-Wide Yeast Fitness Profiling Experiment On Board Orion’s Artemis 1 Mission." 70th International Astronautical Congress (IAC), Washington, DC, October 21-25, 2019. 17. Zea, L., Niederwieser, T., Stodieck, L,. Carr, C., Moeller, R., Nislow, C., Experiment Design for a Genome-Wide Yeast Fitness Profiling Experiment On Board Orion’s Artemis 1 Mission, IAC-19-A2.7.9x51501, 70thInternational Astronautical Congress (IAC) 2019, Washington, D.C.

NOTE: End date changed to 04/29/2024 per NSSC information (Ed., 5/31/23)

NOTE: End date changed to 04/30/2023 per NSSC information (Ed., 4/12/23)

The first aim of this project is to identify the metabolic and genomic pathways in yeast affected by microgravity, space radiation, and a combination of both. The second one is to differentiate between gravity and radiation exposure on single-gene deletion mutants’ ability to thrive in the spaceflight environment. We hypothesize that mutants lacking genes associated with DNA repair, recombination, and replication will have lower survivability rates beyond the van Allen belts than their below van Allen belts- or Earth-controls

The experiment is designed to have a controlled start after Orion is past the van Allen belts, grow ~21 generations of the deletion series, and fix or preserve samples for post-flight analyses. Should the automated controlled approach be considered inappropriate for implementation on EM-1, we have a passive approach that is based on dotting each mutant individually on agar. We have performed both approaches in space in the past.

This project will address three Space Biology Program Science Elements, three Objectives, three Guiding Questions, and four Decadal Survey’s highest priority Recommendations by preserving nucleic acids of different generations of the yeast deletion series cultures grown in space, beyond as well as below the van Allen belts (and uploading the genomic and transcriptomic data to GeneLab).

This aim is complete; we validated the performance of the hardware to support robust cell growth in both the flight and ground control samples. Quantitative assessment of the data was performed using light scattering (at 600nm), and the results indicate no significant difference between flight or ground controls or between sample replicates. This data will be included in the initial biologically focused Deep Space Radiation Genomics (DSRG) manuscript.

2. Assessment of cell viability

Cell viability was assessed using several complimentary analyses, including spot essays, and phase contrast microscopy hemocytometry, and cell morphology was assessed using scanning electron microscopy (SEM); see below.

3. DNA barcode enumeration for fitness profiling

A primary aim of this project was to ask if any particular deletion or overexpression strains became enriched or depleted during the course of growth in the flight and ground samples. Briefly this is accomplished by extracting genomic DNA from pooled cell samples, amplifying the molecular barcodes that mark each unique strain and performing Next Generation Sequencing (NGS) on the amplified barcodes to rank order all strains according to their abundance. We were surprised to discover that the genomic DNA templates from both the flight and ground samples appeared to be damaged, i.e., the polymerase chain reaction (PCR) amplification of the barcodes yielded inconsistent barcode counts using NGS. We therefore attempted to repair the genomic DNA using a panel of enzymes and DNA cleanup techniques that have previously been used on ancient DNA samples. Our systematic evaluation and optimization of the DNA repair procedure took several months. We have treated both flight and ground samples and subjected them to additional rounds of sequencing, and the preliminary results look promising.

4. RNA integrity analysis for transcriptome analysis

RNA extracted from both flight and ground samples has been assessed by standard measures, including bioanalyzer and gel electrophoresis. The transcriptomic analysis of these samples is underway using both short-read NGS and Oxford nanopore single molecule sequencing. We expect this aim to be complete by May 2024.

NOTE: End date changed to 04/30/2023 per NSSC information (Ed., 4/12/23)

The first aim of this project is to identify the metabolic and genomic pathways in yeast affected by microgravity, space radiation, and a combination of both. The second one is to differentiate between gravity and radiation exposure on single-gene deletion mutants’ ability to thrive in the spaceflight environment. We hypothesize that mutants lacking genes associated with DNA repair, recombination, and replication will have lower survivability rates beyond the van Allen belts than their below van Allen belts- or Earth-controls

The experiment is designed to have a controlled start after Orion is past the van Allen belts, grow ~21 generations of the deletion series, and fix or preserve samples for post-flight analyses. Should the automated controlled approach be considered inappropriate for implementation on EM-1, we have a passive approach that is based on dotting each mutant individually on agar. We have performed both approaches in space in the past.

This project will address three Space Biology Program Science Elements, three Objectives, three Guiding Questions, and four Decadal Survey’s highest priority Recommendations by preserving nucleic acids of different generations of the yeast deletion series cultures grown in space, beyond as well as below the van Allen belts (and uploading the genomic and transcriptomic data to GeneLab).

The first aim of this project is to identify the metabolic and genomic pathways in yeast affected by microgravity, space radiation, and a combination of both. The second one is to differentiate between gravity and radiation exposure on single-gene deletion mutants’ ability to thrive in the spaceflight environment. We hypothesize that mutants lacking genes associated with DNA repair, recombination, and replication will have lower survivability rates beyond the van Allen belts than their below van Allen belts- or Earth-controls

The experiment is designed to have a controlled start after Orion is past the van Allen belts, grow ~21 generations of the deletion series, and fix or preserve samples for post-flight analyses. Should the automated controlled approach be considered inappropriate for implementation on EM-1, we have a passive approach that is based on dotting each mutant individually on agar. We have performed both approaches in space in the past.

This project will address three Space Biology Program Science Elements, three Objectives, three Guiding Questions, and four Decadal Survey’s highest priority Recommendations by preserving nucleic acids of different generations of the yeast deletion series cultures grown in space, beyond as well as below the van Allen belts (and uploading the genomic and transcriptomic data to GeneLab).

2. All PLASM units tested under Artemis I-like mission profile for further battery characterization.

3. The Deep Space Radiation Genomics (DSRG) Experiment Verification Test (EVT) results were presented in three conferences; our code to determine Orion’s position with respect to Earth’s magnetosphere was released in GitHub for open access; and a book was published, partly supported by this grant.

4. SRFR with NASA Headquarters (HQ) passed and team is ready for Space Launch System (SLS) launch.

The first aim of this project is to identify the metabolic and genomic pathways in yeast affected by microgravity, space radiation, and a combination of both. The second one is to differentiate between gravity and radiation exposure on single-gene deletion mutants’ ability to thrive in the spaceflight environment. We hypothesize that mutants lacking genes associated with DNA repair, recombination, and replication will have lower survivability rates beyond the van Allen belts than their below van Allen belts- or Earth-controls

The experiment is designed to have a controlled start after Orion is past the van Allen belts, grow ~21 generations of the deletion series, and fix or preserve samples for post-flight analyses. Should the automated controlled approach be considered inappropriate for implementation on EM-1, we have a passive approach that is based on dotting each mutant individually on agar. We have performed both approaches in space in the past.

This project will address three Space Biology Program Science Elements, three Objectives, three Guiding Questions, and four Decadal Survey’s highest priority Recommendations by preserving nucleic acids of different generations of the yeast deletion series cultures grown in space, beyond as well as below the van Allen belts (and uploading the genomic and transcriptomic data to GeneLab).

2. A new hardware subsystem for temperature control was designed, developed, tested, and integrated

3. The Deep Space Radiation Genomics (DSRG) Experiment Verification Test (EVT) was completed, yielding “excellent” score under all of the mission success criteria

Other hardware and protocol work performed during this project’s second year include:

• Peristaltic pump characterization (flow and power consumption as function of operational duration)

• Characterization of metabolic pressure inside Culture Bags as function of time and environmental temperature

• Optimization of the Culture Bag sealing procedure

• Optimization of the Fluidic System assembly, sterilization, loading, and acceptance testing procedure

• Data production for Safety Review 0/I/II

• Production of all structural parts for all five Peristaltic Laboratory for Automated Science with Multigenerations (PLASM) units

• HFIT buy-in for the ISS units

• Flight-like software development

• Integrated software and accelerometer testing to verify our software architecture can detect launch and autonomously activate the experiment passed the Van Allen belts

• Development of a python code yielding a timetable to let PLASM know when can it activate the experiment, to ensure yeast growth occurs outside of Earth’s magnetosphere, hence exposing the cells to the most pristine deep space radiation environment possible (i.e., not confounded by our magnetosphere).

The first aim of this project is to identify the metabolic and genomic pathways in yeast affected by microgravity, space radiation, and a combination of both. The second one is to differentiate between gravity and radiation exposure on single-gene deletion mutants’ ability to thrive in the spaceflight environment. We hypothesize that mutants lacking genes associated with DNA repair, recombination, and replication will have lower survivability rates beyond the van Allen belts than their below van Allen belts- or Earth-controls

The experiment is designed to have a controlled start after Orion is past the van Allen belts, grow ~21 generations of the deletion series, and fix or preserve samples for post-flight analyses. Should the automated controlled approach be considered inappropriate for implementation on EM-1, we have a passive approach that is based on dotting each mutant individually on agar. We have performed both approaches in space in the past.

This project will address three Space Biology Program Science Elements, three Objectives, three Guiding Questions, and four Decadal Survey’s highest priority Recommendations by preserving nucleic acids of different generations of the yeast deletion series cultures grown in space, beyond as well as below the van Allen belts (and uploading the genomic and transcriptomic data to GeneLab).

2. Two irradiation campaigns have taken place to provide data on what may we expect from the Artemis 1 mission, further verifying the validity of our experimental design. Additionally, a 69-day long Science Verification Test (SVT) is currently ongoing and already showing effective yeast growth in flight-like hardware.

The first aim of this project is to identify the metabolic and genomic pathways in yeast affected by microgravity, space radiation, and a combination of both. The second one is to differentiate between gravity and radiation exposure on single-gene deletion mutants’ ability to thrive in the spaceflight environment. We hypothesize that mutants lacking genes associated with DNA repair, recombination, and replication will have lower survivability rates beyond the van Allen belts than their below van Allen belts- or Earth-controls

The experiment is designed to have a controlled start after Orion is past the van Allen belts, grow ~21 generations of the deletion series, and fix or preserve samples for post-flight analyses. Should the automated controlled approach be considered inappropriate for implementation on EM-1, we have a passive approach that is based on dotting each mutant individually on agar. We have performed both approaches in space in the past.

This project will address three Space Biology Program Science Elements, three Objectives, three Guiding Questions, and four Decadal Survey’s highest priority Recommendations by preserving nucleic acids of different generations of the yeast deletion series cultures grown in space, beyond as well as below the van Allen belts (and uploading the genomic and transcriptomic data to GeneLab).