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Project Title:  Using Water Bears to Identify Biological Countermeasures to Stress During Multigenerational Spaceflight Reduce
Images: icon  Fiscal Year: FY 2019 
Division: Space Biology 
Research Discipline/Element:
Space Biology: Cell & Molecular Biology   | Animal Biology: Invertebrate  
Start Date: 11/01/2014  
End Date: 08/26/2019  
Task Last Updated: 08/30/2019 
Download report in PDF pdf
Principal Investigator/Affiliation:   Boothby, Thomas  Ph.D. / University of Wyoming 
Address:  1000 E. University Ave., Department #3944 
 
Laramie , WY 82071 
Email: Thomas.Boothby@uwyo.edu 
Phone:   
Congressional District:
Web:  
Organization Type: UNIVERSITY 
Organization Name: University of Wyoming 
Joint Agency:  
Comments: NOTE: Previously at University of North Carolina until fall 2019.  
Co-Investigator(s)
Affiliation: 
Goldstein, Bob  Ph.D. University of North Carolina 
Key Personnel Changes / Previous PI: September 2016 report: Kiera Patanella, an undergraduate at the University of North Carolina at Chapel Hill working on this project, has graduated and obtained her bachelors degree in Biology. Cody Weyhrich, an undergraduate at the University of North Carolina at Chapel Hill, has started working on this project as of 8/1/2016.
Project Information: Grant/Contract No. NNX15AB44G 
Responsible Center: NASA ARC 
Grant Monitor: Sato, Kevin  
Center Contact: 650-604-1104 
kevin.y.sato@nasa.gov 
Solicitation: 2014 Space Biology Flight NNH14ZTT001N 
Grant/Contract No.: NNX15AB44G 
Project Type: FLIGHT 
Flight Program: ISS 
No. of Post Docs:
No. of PhD Candidates:  
No. of Master's Candidates:  
No. of Bachelor's Candidates:
No. of PhD Degrees:  
No. of Master's Degrees:  
No. of Bachelor's Degrees:  
Space Biology Element: (1) Cell & Molecular Biology
(2) Animal Biology: Invertebrate
Space Biology Cross-Element Discipline: (1) Reproductive Biology
(2) Developmental Biology
Space Biology Special Category: None
Flight Assignment/Project Notes: NOTE: End date changed to 8/26/2019 due to PI move to University of Wyoming; new grant 80NSSC20K0283 awarded (Ed., 7/24/2020)

NOTE: Extended to 9/02/2020 per F. Hernandez/ARC (Ed., 10/12/18)

NOTE: Extended to 10/31/2018 per F. Hernandez/ARC (Ed., 12/6/17)

Task Description: For most organisms the stresses associated with spaceflight induce a variety of detrimental effects. To foster a safe and productive long-term human presence in space, therapies and countermeasures to spaceflight-induced stress should be developed. Tardigrades (water bears) are polyextremophiles that have evolved to tolerate multiple extreme environments, which are restrictive to most life. In 2007 tardigrades were shown to survive and reproduce normally during an 11-day low Earth orbit on the Foton-M3 Capsule. We speculate that mechanisms tardigrades have evolved to withstand extreme environments on Earth may, as a side-effect, confer protection against the stresses of spaceflight. This makes tardigrades a uniquely valuable system for studying responses to spaceflight. We have sequenced the genome of the tardigrades Hypsibius dujardini, as well as developed and validated experimental and computational approaches for measuring the effect of different environmental conditions on tardigrade gene expression – allowing us to identify mechanisms used by tardigrades to protect themselves from different stresses. We have also developed a reverse genetic approach, RNA interference, for tardigrades that allows us to directly investigate the role of a gene in conferring tolerance to an environment. We will use these approaches to study tardigrades’ initial, as well as multigenerational, response to spaceflight and use RNA interference to test the functionality of the genes identified in our study. Next-generation transcriptome sequencing will be conducted on tardigrades cultures kept 0 generations (founding generation) and 4 generations onboard the International Space Station (ISS). Differential expression analysis will be conducted to compare ISS spaceflight timepoints, ground controls, and tardigrades exposed to other extreme stresses (e.g., desiccation, freezing). This approach will allow us to identify potential mediators of stress tolerance, which will serve as candidates for functional RNA interference experiments. Understanding how tardigrades tolerate spaceflight will better guide future research into countermeasures and therapies for humans exposed to the stresses of prolonged space travel. This proposal's strengths are: the use of an organism that is suited to studying mechanisms of multigenerational tolerance of extreme environments and that has an established RNA interference method for confirming the function of genes identified in our study, our Preliminary Results that validate our proposed approach and technical capabilities as well as the uniqueness and suitability of tardigrades that will allow us to conduct this study. The participants for this study are comprised of experts in tardigrades' stress response and have considerable experience with next-generation sequencing and analysis of non-model organisms. The proposed experiments directly address recommendation AH16 of the Decadal Survey and are in line with recommendation OCB-5 and CMM-5 of NASA’s Multigenerational and Developmental Biology of Invertebrates Research Emphasis as well as NASA’s Fundamental Space Biology Plan 2010-2020 goals. Completion of our proposal will identify genes required for tardigrades to survive multigenerational spaceflight and will be a key step towards developing countermeasures and therapies for stresses associated with prolonged human exposure to space environments.

Research Impact/Earth Benefits: Along with using mechanisms of stress tolerance to counteract detrimental effects of space travel, data from our proposed experiments could be used in the long term toward solving serious problems in the field of human health. Utilizing mechanisms that allow tardigrades to stabilize their cellular proteins and nucleic acids has been proposed as an option for the dry storage and stabilization of vaccines and other biomaterials (Guo et al., 2000; Wolkers et al., 2001; Puhlev et al., 2001). Because current techniques for vaccine production, distribution, and storage nearly always require a constant cold chain (e.g., -80 and 20 degrees C freezers), these processes are extremely expensive. Some estimates put cold chain costs at around 80% of the total cost of vaccination (Chen et al., 2011). By generating additional stress response datasets, such as response to microgravity, freezing, irradiation, and hypoxia, we will increase our ability and that of other researchers to identify specific mediators of desiccation tolerance, which will then be applied to this and similar problems.

Additionally, a better understanding of mechanisms of stress tolerance could lead to the development of drought and/or freeze tolerant crops.

Guo, N., Puhlev, I., Brown, D. R., Mansbridge, J., & Levine, F. (2000). Trehalose expression confers desiccation tolerance on human cells. Nature biotechnology, 18(2), 168-171.

Wolkers, W. F., Walker, N. J., Tablin, F., & Crowe, J. H. (2001). Human platelets loaded with trehalose survive freeze-drying. Cryobiology, 42(2), 79-87.

Puhlev, I., Guo, N., Brown, D. R., & Levine, F. (2001). Desiccation tolerance in human cells. Cryobiology, 42(3), 207-217.

Chen, X. et al. (2011). Improving the reach of vaccines to low-resource regions, with a needle-free vaccine delivery device and long-term thermostabilization. J. Controlled Release 152, 349–355.

Task Progress & Bibliography Information FY2019 
Task Progress: REPORTING FROM AUGUST 2019; PRINCIPAL INVESTIGATOR MOVED TO UNIVERSITY OF WYOMING IN FALL 2019 and NEW GRANT 80NSSC20K0283 AWARDED (Ed., 7/24/2020)

The bulk of our research on NASA Grant NNX15AB44G that was conducted at UNC (the University of North Carolina) can be broken down into two categories: 1.) ground controls and experiments, and 2.) testing and validation for our flight experiment. Below are summaries of our research in both of these categories. Ground Controls and Experiments

Identification and functional assays to identify mediators of tardigrade desiccation tolerance: To prepare for comparison of changes in gene expression manifested by tardigrades (water bears) exposed to multigenerational spaceflight with ground-based stresses we have begun gathering transcriptomic datasets from terrestrial stresses. In addition we have performed functional experiments to assess which candidate genes from our transcriptomic datasets are functional mediators of desiccation (drying) tolerance.

Our major findings from these endeavors have been published in Boothby et al., 2017 (Boothby TC, Pielak GJ. Intrinsically disordered proteins and desiccation tolerance: Elucidating functional and mechanistic underpinnings of anhydrobiosis. Bioessays. 2017 Nov;39(11):700119. Epub 2017 Sep 13. https://doi.org/10.1002/bies.201700119 ; PubMed PMID: 28901557 ; Ed. Note 9/9/19: reported in August 2018 FY2018 Task Book report Bibliography ). These results will be summarized here briefly since detailed descriptions of the experiments, methods, and results are presented in publication.

To identify genes that might play a role in tardigrade desiccation tolerance, we extracted and sequenced RNA from tardigrades that had either been left unstressed in culture or desiccated. Comparison of transcript levels coming from each predicted gene was conducted and genes ranked based on fold change (how much expression of the gene increased during drying) and overall abundance (how many transcripts per million transcripts were coming from a particular gene).

The main takeaway from this comparison was that a class of tardigrade specific genes known as Cytosolic Abundant Heat Soluble (CAHS) genes are upregulated heavily during desiccation. We performed RNA interference experiments in tardigrades to reduce the level of expression of these genes and found that the animals no longer robustly survived drying when CAHS genes were targeted. We also found that expressing these genes in bacteria and yeast (which normally do not have these genes) led to up to two orders of magnitude increases (100X) in desiccation tolerance. Amazingly, when purified CAHS proteins were found to protect biological material (the enzyme lactate dehydrogenase) about an order of magnitude (10X) better than current FDA approved excipient trehalose and serum albumin.

Finally, we correlate the protective capabilities of these CAHS proteins to their ability to form vitrified (glass-like) solids, as opposed to crystalline solids. These finding may be of interest to NASA, as this presents an avenue for stabilizing and protecting biological material in a dry state without refrigeration. This might be useful for prolonged storage of biomaterials on the ISS or other spaceflight missions where freezer and refrigeration space is limited or logistically difficult.

Exploring cross-tolerance between desiccation and freeze tolerances in tardigrades:

Tardigrades survive an amazing number of abiotic stresses, and in some cases the severity of these stresses is well beyond that tardigrades would ever experience in nature (e.g., temperatures close to absolute zero, thousands of gray of radiation, the vacuum of outer space). The question therefore arises--how did tardigrades evolve tolerance to stresses they have never experienced? One hypothesis is that as tardigrades moved onto land from the ocean (where they originally evolved) they developed desiccation tolerance in response to their new, dryer, conditions and as a by-product became tolerant to other stresses. If this hypothesis is correct, then the mediators that tardigrades use to survive desiccation should in theory be the same mediators they use to survive other stresses. To assess if this is true we performed transcriptome sequencing on tardigrades that had been frozen, and compared this data to our previous datasets (desiccated and unstressed).

Surprisingly, we found that changes in gene expression between desiccated and frozen tardigrades are highly divergent. In fact, either stress condition is more similar to unstressed conditions than they are to each other. Most telling, we observed that expression of CAHS genes was not influenced by freezing conditions, and furthermore RNA interference targeting these genes did not result in statistical decreases in survival in tardigrades exposed to freezing conditions.

These results are presented in detail in Boothby et al., 2017 (Boothby TC, Pielak GJ. Intrinsically disordered proteins and desiccation tolerance: Elucidating functional and mechanistic underpinnings of anhydrobiosis. Bioessays. 2017 Nov;39(11):700119. Epub 2017 Sep 13. https://doi.org/10.1002/bies.201700119 ; PubMed PMID: 28901557 ; Ed. Note 9/9/19: reported in August 2018 FY2018 Task Book report Bibliography ).

We are now delving more into our frozen transcriptome to identify functional mediators of freeze tolerance in tardigrades using a similar approach to the one taken for our desiccation study.

Understanding to commonalities and differences between how tardigrades survive freezing and desiccation is an important facet of our overall strategy for this project, as spaceflight induced changes in gene response will ultimately be compared to ground-based stress responses. Comparing changes in gene expression for ground-based stresses now will help us understand the overlap with spaceflight induced changes later.

How do tardigrade CAHS proteins mediate desiccation tolerance?

To better understand of CAHS proteins protect tardigrades and other biological material and cells from the harmful effects of drying, and how these proteins might protect biological materials from other stresses (included spaceflight), we have been characterizing the biochemical and biophysical nature of these proteins.

We have discovered that these proteins behave in a very peculiar way. At room temperature and at concentrations greater than or equal to 30 g/L these proteins form reversible hydrogels. We have characterized the gel state of these proteins via cone plate rheometry as well as scanning electron microscopy. Both techniques clearly demonstrate that these proteins have classic gel-like behavior and morphology. In retrospect it makes sense that these proteins form hydrogels, as hydrogels are known to form vitrified solids when dried (see above).

We were curious if the gel state of these proteins is important for their protective capabilities. To probe this, we used 19-F NMR to look at the folded state of a test protein, SH3. SH3 is a ‘metastable’ protein, meaning that normally (in solution) SH3 is in a folded state ~50% of the time and in an unfolded state about ~50% of the time. This is easily measured using 19-F NMR. We first tested SH3 in solution, and as expected two clear peaks (folded and unfolded) were present. We then co-incubated SH3 with a CAHS protein (at increasing concentrations) and looked at the levels of folded and unfolded protein. We found that CAHS proteins had no noticeable effect on SH3 folding below 30 g/L. However, above 30 g/L of CAHS protein, there was a reduction in the level of unfolded SH3 protein, and a corresponding increase in folded SH3. Interestingly, 30 g/L is the concentration determined by cone plate rheometer as the gel point for these proteins. The hydrogels that CAHS proteins form are reverse and heat dependent. Therefore, we tested whether heating to induce the breakdown of the hydrogel influenced SH3 folding. We found when heated from 19C to 42C the CAHS proteins went back into solution (the gel state vanished) and there was a return of an SH3 unfolded population. Cooling this solution back down to 19C resulted in re-gelling of the CAHS proteins and a corresponding disappearance of the SH3 unfolded species and an increase in the SH3 folded species. Therefore, it appears that there is a strong correlation between the gelled state of CAHS proteins and their ability to stabilize proteins in a folded state.

We are now characterizing the sequence features of CAHS proteins at allow them to form gels. These studies are being conducted by making mutant versions of the proteins and testing their ability to form gels and protect biological materials.

Testing and Validation for Flight Experiment

The bulk of this effort has been made in preparing for and performing our Science Validation Test (SVT-1).

The main goal of SVT-1 was to compare the efficacy of culturing tardigrades (Hypsibius exemplasris) using the CellMax and PI Start Kit (PISK).

Syringes containing ~500 tardigrades or concentrated algae were prepared using the same stock cultures. Syringes were frozen at -80 degrees Celsius and either stored at this temperature or shipped on dry ice to NASA Ames.

On May 10th, tardigrades were injected into 3 PISK bioreactors (at NASA Ames) and 3 CellMax Bioreactors (at UNC).

Temperature control for PISK bioreactors was carried out using integrated temperature control – which varied between ~15 – 17 degrees Celsius. Oxygenation for PISK bioreactors was carried out via gas dosing using medical grade air.

Temperature control for CellMax bioreactors was carried out by placing the CellMax system in a controlled temperature chamber. The initial temperature used was 15 degrees Celsius, but was changed to 16 degrees Celsius to more closely mirror PISK temperatures. Oxygenation was not controlled, but rather relied on the passive transport of oxygen across the CellMax systems permeable tubing.

Every 7 days, up to day 21, subsamples (300 ul) were extracted from both PISK and CellMax bioreactors using syringes. These samples were stored at -80 degrees Celsius. At day 14, fresh algae (food source) was injected into each PISK and CellMax bioreactor. At day 28, the experiment ended and whole bioreactors were detached and frozen at -80 degrees Celsius. PISK subsample syringes and bioreactors were returned frozen on dry ice to the Principal Investigator's (PI) lab at UNC.

Upon receipt at the PI’s lab, samples were transferred from their shipping unit to a -80 degree freezer.

To compare the viability of culturing using the PISK, all samples were thawed. For each subsample (300 ul) the entire volume of thawed sample was analyzed by direct observation using a dissecting microscope. Total animal counts were taken and densities calculated. Similarly, for bioreactors, the entire contents of the bioreactor was thawed and transferred to a 15 mL tube. Three 50 ul samples were taken from each tube and total animal counts made and densities calculated.

The remaining bioreactor contents was fixed with RNAlater and processed for total RNA extraction.

Tardigrade Densities and Total Counts

Prior to testing our definition for success was achieving Day 28 animal densities in the PISK bioreactors that were above or within 20% of the densities from our CellMax bioreactors. Our average PISK animal density at Day 28 was 357.8 animals/mL where as our CellMax Day 28 animal density was slightly lower at 282.2 animals/mL. By our original definition, the PISK successfully competed with the CellMax system with regards to effectively culturing tardigrades over a 28 day period.

REPORTING IN AUGUST 2018

We have been preparing for our flight experiment. This has mostly manifested itself in the development of an SRD (Science Requirements Document), protocols, and coordinating with the engineering team who are doing some final tests on our bioreactor setup. Our next step will be to actually get our animals in the hardware and do ground tests.

Bibliography Type: Description: (Last Updated: 07/24/2020)  Show Cumulative Bibliography Listing
 
Abstracts for Journals and Proceedings Boothby TC, Piszkiewicz S, Mehta A, Brozena A, Tapia H, Koshland D, Holehouse A, Pappu R, Goldstein B, Pielak G. "Gelation and Vitrification of Tardigrade IDPs." 62nd Annual Meeting of the Biophysical Society, San Francisco, CA, February 17-21, 2018.

Biophysical Journal. 2018 Feb 2;114(3 Suppl):560a-561a. https://doi.org/10.1016/j.bpj.2017.11.3065 , Feb-2018

Articles in Peer-reviewed Journals Piszkiewicz S, Gunn KH, Warmuth O, Propst A, Mehta A, Nguyen KH, Kuhlman E, Guseman AJ, Stadmiller SS, Boothby TC, Neher SB, Pielak GJ. "Protecting activity of desiccated enzymes." Protein Sci. 2019 May;28(5):941-51. Epub 2019 Mar 30. https://doi.org/10.1002/pro.3604 ; PubMed PMID: 30868674; PubMed Central PMCID: PMC6459994 , May-2019
Articles in Peer-reviewed Journals Boothby TC. "Desiccation of Hypsibius exemplaris." Cold Spring Harb Protoc. 2018 Nov;18(11):871-3. https://doi.org/10.1101/pdb.prot102327 ; PubMed PMID: 30385670 , Nov-2018
Articles in Peer-reviewed Journals Boothby TC. "Total RNA extraction from tardigrades." Cold Spring Harb Protoc. 2018 Nov 1;2018(11):905-7. https://doi.org/10.1101/pdb.prot102376 ; PubMed PMID: 30385675 , Nov-2018
Articles in Peer-reviewed Journals Boothby TC, Pielak GJ. "Intrinsically disordered proteins and desiccation tolerance: Elucidating functional and mechanistic underpinnings of anhydrobiosis." Bioessays. 2017 Nov;39(11):700119. Epub 2017 Sep 13. https://doi.org/10.1002/bies.201700119 ; PubMed PMID: 28901557 , Nov-2017
Articles in Peer-reviewed Journals Boothby TC. "Mechanisms and evolution of resistance to environmental extremes in animals." Evodevo. 2019 Nov 18;10:30. Review. https://doi.org/10.1186/s13227-019-0143-4 ; PMID: 31827759; PMCID: PMC6862762 , Nov-2019
Journal/Magazine covers Piszkiewicz S, Gunn KH, Warmuth O, Propst A, Mehta A, Nguyen KH, Kuhlman E, Guseman AJ, Stadmiller SS, Boothby TC, Neher SB, Pielak GJ. "Cover in journal Protein Science for article, 'Protecting activity of desiccated enzymes.' " Protein Sci. 2019 May;28(5):941-51. https://onlinelibrary.wiley.com/doi/abs/10.1002/pro.3465 ; PubMed PMID: 30868674 , May-2019
Project Title:  Using Water Bears to Identify Biological Countermeasures to Stress During Multigenerational Spaceflight Reduce
Images: icon  Fiscal Year: FY 2018 
Division: Space Biology 
Research Discipline/Element:
Space Biology: Cell & Molecular Biology   | Animal Biology: Invertebrate  
Start Date: 11/01/2014  
End Date: 10/31/2018  
Task Last Updated: 09/04/2017 
Download report in PDF pdf
Principal Investigator/Affiliation:   Boothby, Thomas  Ph.D. / University of Wyoming 
Address:  1000 E. University Ave., Department #3944 
 
Laramie , WY 82071 
Email: Thomas.Boothby@uwyo.edu 
Phone:   
Congressional District:
Web:  
Organization Type: UNIVERSITY 
Organization Name: University of Wyoming 
Joint Agency:  
Comments: NOTE: Previously at University of North Carolina until fall 2019.  
Co-Investigator(s)
Affiliation: 
Goldstein, Bob  Ph.D. University of North Carolina 
Key Personnel Changes / Previous PI: September 2016 report: Kiera Patanella, an undergraduate at the University of North Carolina at Chapel Hill working on this project, has graduated and obtained her bachelors degree in Biology. Cody Weyhrich, an undergraduate at the University of North Carolina at Chapel Hill, has started working on this project as of 8/1/2016.
Project Information: Grant/Contract No. NNX15AB44G 
Responsible Center: NASA ARC 
Grant Monitor: Sato, Kevin  
Center Contact: 650-604-1104 
kevin.y.sato@nasa.gov 
Solicitation: 2014 Space Biology Flight NNH14ZTT001N 
Grant/Contract No.: NNX15AB44G 
Project Type: FLIGHT 
Flight Program: ISS 
No. of Post Docs:
No. of PhD Candidates:  
No. of Master's Candidates:  
No. of Bachelor's Candidates:
No. of PhD Degrees:  
No. of Master's Degrees:  
No. of Bachelor's Degrees:
Space Biology Element: (1) Cell & Molecular Biology
(2) Animal Biology: Invertebrate
Space Biology Cross-Element Discipline: (1) Reproductive Biology
(2) Developmental Biology
Space Biology Special Category: None
Flight Assignment/Project Notes: NOTE: Extended to 10/31/2018 per F. Hernandez/ARC (Ed., 12/6/17)

Task Description: For most organisms the stresses associated with spaceflight induce a variety of detrimental effects. To foster a safe and productive long-term human presence in space, therapies and countermeasures to spaceflight-induced stress should be developed. Tardigrades (water bears) are polyextremophiles that have evolved to tolerate multiple extreme environments, which are restrictive to most life. In 2007 tardigrades were shown to survive and reproduce normally during an 11-day low Earth orbit on the Foton-M3 Capsule. We speculate that mechanisms tardigrades have evolved to withstand extreme environments on Earth may, as a side-effect, confer protection against the stresses of spaceflight. This makes tardigrades a uniquely valuable system for studying responses to spaceflight. We have sequenced the genome of the tardigrades Hypsibius dujardini, as well as developed and validated experimental and computational approaches for measuring the effect of different environmental conditions on tardigrade gene expression – allowing us to identify mechanisms used by tardigrades to protect themselves from different stresses. We have also developed a reverse genetic approach, RNA interference, for tardigrades that allows us to directly investigate the role of a gene in conferring tolerance to an environment. We will use these approaches to study tardigrades’ initial, as well as multigenerational response to spaceflight and use RNA interference to test the functionality of the genes identified in our study. Next-generation transcriptome sequencing will be conducted on tardigrades cultures kept 0 generations (founding generation) and 4 generations onboard the International Space Station (ISS). Differential expression analysis will be conducted to compare ISS spaceflight timepoints, ground controls, and tardigrades exposed to other extreme stresses (e.g., desiccation, freezing). This approach will allow us to identify potential mediators of stress tolerance, which will serve as candidates for functional RNA interference experiments. Understanding how tardigrades tolerate spaceflight will better guide future research into countermeasures and therapies for humans exposed to the stresses of prolonged space travel. This proposal's strengths are: the use of an organism that is suited to studying mechanisms of multigenerational tolerance of extreme environments and that has an established RNA interference method for confirming the function of genes identified in our study, our Preliminary Results that validate our proposed approach and technical capabilities as well as the uniqueness and suitability of tardigrades that will allow us to conduct this study. The participants for this study are comprised of experts in tardigrades' stress response and have considerable experience with next-generation sequencing and analysis of non-model organisms. The proposed experiments directly address recommendation AH16 of the Decadal Survey and are in line with recommendation OCB-5 and CMM-5 of NASA’s Multigenerational and Developmental Biology of Invertebrates Research Emphasis as well as NASA’s Fundamental Space Biology Plan 2010-2020 goals. Completion of our proposal will identify genes required for tardigrades to survive multigenerational spaceflight and will be a key step towards developing countermeasures and therapies for stresses associated with prolonged human exposure to space environments.

Research Impact/Earth Benefits: Along with using mechanisms of stress tolerance to counteract detrimental effects of space travel, data from our proposed experiments could be used in the long term toward solving serious problems in the field of human health. Utilizing mechanisms that allow tardigrades to stabilize their cellular proteins and nucleic acids has been proposed as an option for the dry storage and stabilization of vaccines and other biomaterials (Guo et al., 2000; Wolkers et al., 2001; Puhlev et al., 2001). Because current techniques for vaccine production, distribution, and storage nearly always require a constant cold chain (e.g., -80 and 20 degrees C freezers), these processes are extremely expensive. Some estimates put cold chain costs at around 80% of the total cost of vaccination (Chen et al., 2011). By generating additional stress response datasets, such as response to microgravity, freezing, irradiation, and hypoxia, we will increase our ability and that of other researchers to identify specific mediators of desiccation tolerance, which will then be applied to this and similar problems.

Additionally, a better understanding of mechanisms of stress tolerance could lead to the development of drought and/or freeze tolerant crops.

Guo, N., Puhlev, I., Brown, D. R., Mansbridge, J., & Levine, F. (2000). Trehalose expression confers desiccation tolerance on human cells. Nature biotechnology, 18(2), 168-171.

Wolkers, W. F., Walker, N. J., Tablin, F., & Crowe, J. H. (2001). Human platelets loaded with trehalose survive freeze-drying. Cryobiology, 42(2), 79-87.

Puhlev, I., Guo, N., Brown, D. R., & Levine, F. (2001). Desiccation tolerance in human cells. Cryobiology, 42(3), 207-217.

Chen, X. et al. (2011). Improving the reach of vaccines to low-resource regions, with a needle-free vaccine delivery device and long-term thermostabilization. J. Controlled Release 152, 349–355.

Task Progress & Bibliography Information FY2018 
Task Progress: Work/progress on this project can be generally broken down into two categories-- 1.) preparation and hardware testing for our flight mission, and 2.) execution of complimentary ground experiments.

1.) Preparation for flight experiments. Initially we selected the BIOS culture system of our flight experiments and conducted extensive ground testing to verify all parameters of our flight experiment with this hardware. Since the wait time for the use of the BIOS culture system is significant we have been exploring additional hardware options, which at the time we believed might help us accelerate the timing of our flight experiment. We have been in contact with BioServe and Techshot regarding different hardware options and were validating use of BioServe's BioCell. This hardware seems to work well for our purposes, but apparently new information on launch schedules has come out and using this hardware will no longer accelerate our project's expected launch date. Thus, we have settled on using our original hardware choice, the BIOS culture system.

2.) Complimentary ground experiments. Part of our post-flight analysis will be to compare the genes unregulated during exposure to the stresses of spaceflight with ground based stresses. To this end we have been studying the change in gene expression in tardigrades exposed to different Earth-based stresses. Our study of tardigrade responses to drying has resulted in a paper published in Molecular Cell (Boothby et al., 2017). We are currently preparing a manuscript looking at the connection (or lack of connection) between drying and freezing tolerance in tardigrades.

Bibliography Type: Description: (Last Updated: 07/24/2020)  Show Cumulative Bibliography Listing
 
Abstracts for Journals and Proceedings Boothby TC, Piszkiewicz S, Mehta A, Brozena A, Tapia H, Koshland D, Holehouse A, Pappu R, Goldstein B, Pielak G. "Tardigrade Disordered Proteins Mediate Desiccation Tolerance." Presented at the 61st Annual Meeting of the Biophysical Society, New Orleans, LA, February 11-15, 2017.

Biophysical Journal. 2017 Feb 3;112(3 Suppl):480a. https://doi.org/10.1016/j.bpj.2016.11.2600 , Feb-2017

Abstracts for Journals and Proceedings Boothby TC, Tapia H, Brozena AH, Piszkiewicz S, Smith AE, Mehta A, Koshland D, Goldstein B, Pielak G. "How Do Tardigades Survive Extremes? Disordered Proteins as Mediators of Tardigrade Stress Tolerance." Society for Integrative and Comparative Biology Annual Meeting 2017, New Orleans, LA, January 4-8, 2017.

Integrative and Comparative Biology. 2017 Mar 1;57(Suppl 1):E208. , Mar-2017

Articles in Peer-reviewed Journals Boothby TC, Tapia H, Brozena AH, Piszkiewicz S, Smith AE, Giovannini I, Rebecchi L, Pielak GJ, Koshland D, Goldstein B. "Tardigrades use intrinsically disordered proteins to survive desiccation." Molecular Cell. 2017 Mar 16;65(6):975-84.e5. https://doi.org/10.1016/j.molcel.2017.02.018 ; PubMed PMID: 28306513 , Mar-2017
Articles in Peer-reviewed Journals Boothby TC, Pielak GJ. "Intrinsically disordered proteins and desiccation tolerance: Elucidating functional and mechanistic underpinnings of anhydrobiosis." BioEssays. Version of Record online: 13 SEP 2017. https://doi.org/10.1002/bies.201700119 , Sep-2017
Project Title:  Using Water Bears to Identify Biological Countermeasures to Stress During Multigenerational Spaceflight Reduce
Images: icon  Fiscal Year: FY 2017 
Division: Space Biology 
Research Discipline/Element:
Space Biology: Cell & Molecular Biology   | Animal Biology: Invertebrate  
Start Date: 11/01/2014  
End Date: 10/31/2017  
Task Last Updated: 09/02/2016 
Download report in PDF pdf
Principal Investigator/Affiliation:   Boothby, Thomas  Ph.D. / University of Wyoming 
Address:  1000 E. University Ave., Department #3944 
 
Laramie , WY 82071 
Email: Thomas.Boothby@uwyo.edu 
Phone:   
Congressional District:
Web:  
Organization Type: UNIVERSITY 
Organization Name: University of Wyoming 
Joint Agency:  
Comments: NOTE: Previously at University of North Carolina until fall 2019.  
Co-Investigator(s)
Affiliation: 
Goldstein, Bob  Ph.D. University of North Carolina 
Key Personnel Changes / Previous PI: September 2016 report: Kiera Patanella, an undergraduate at the University of North Carolina at Chapel Hill working on this project, has graduated and obtained her bachelors degree in Biology. Cody Weyhrich, an undergraduate at the University of North Carolina at Chapel Hill, has started working on this project as of 8/1/2016.
Project Information: Grant/Contract No. NNX15AB44G 
Responsible Center: NASA ARC 
Grant Monitor: Sato, Kevin  
Center Contact: 650-604-1104 
kevin.y.sato@nasa.gov 
Solicitation: 2014 Space Biology Flight NNH14ZTT001N 
Grant/Contract No.: NNX15AB44G 
Project Type: FLIGHT 
Flight Program: ISS 
No. of Post Docs:
No. of PhD Candidates:  
No. of Master's Candidates:  
No. of Bachelor's Candidates:
No. of PhD Degrees:  
No. of Master's Degrees:  
No. of Bachelor's Degrees:
Space Biology Element: (1) Cell & Molecular Biology
(2) Animal Biology: Invertebrate
Space Biology Cross-Element Discipline: (1) Reproductive Biology
(2) Developmental Biology
Space Biology Special Category: None
Task Description: For most organisms the stresses associated with spaceflight induce a variety of detrimental effects. To foster a safe and productive long-term human presence in space, therapies and countermeasures to spaceflight-induced stress should be developed. Tardigrades (water bears) are polyextremophiles that have evolved to tolerate multiple extreme environments, which are restrictive to most life. In 2007 tardigrades were shown to survive and reproduce normally during an 11-day low Earth orbit on the Foton-M3 Capsule. We speculate that mechanisms tardigrades have evolved to withstand extreme environments on Earth may, as a side-effect, confer protection against the stresses of spaceflight. This makes tardigrades a uniquely valuable system for studying responses to spaceflight. We have sequenced the genome of the tardigrades Hypsibius dujardini, as well as developed and validated experimental and computational approaches for measuring the effect of different environmental conditions on tardigrade gene expression – allowing us to identify mechanisms used by tardigrades to protect themselves from different stresses. We have also developed a reverse genetic approach, RNA interference, for tardigrades that allows us to directly investigate the role of a gene in conferring tolerance to an environment. We will use these approaches to study tardigrades’ initial, as well as multigenerational response to spaceflight and use RNA interference to test the functionality of the genes identified in our study. Next-generation transcriptome sequencing will be conducted on tardigrades cultures kept 0 generations (founding generation) and 4 generations onboard the International Space Station (ISS). Differential expression analysis will be conducted to compare ISS spaceflight timepoints, ground controls, and tardigrades exposed to other extreme stresses (e.g., desiccation, freezing). This approach will allow us to identify potential mediators of stress tolerance, which will serve as candidates for functional RNA interference experiments. Understanding how tardigrades tolerate spaceflight will better guide future research into countermeasures and therapies for humans exposed to the stresses of prolonged space travel. This proposal's strengths are: the use of an organism that is suited to studying mechanisms of multigenerational tolerance of extreme environments and that has an established RNA interference method for confirming the function of genes identified in our study, our Preliminary Results that validate our proposed approach and technical capabilities as well as the uniqueness and suitability of tardigrades that will allow us to conduct this study. The participants for this study are comprised of experts in tardigrades' stress response and have considerable experience with next-generation sequencing and analysis of non-model organisms. The proposed experiments directly address recommendation AH16 of the Decadal Survey and are in line with recommendation OCB-5 and CMM-5 of NASA’s Multigenerational and Developmental Biology of Invertebrates Research Emphasis as well as NASA’s Fundamental Space Biology Plan 2010-2020 goals. Completion of our proposal will identify genes required for tardigrades to survive multigenerational spaceflight and will be a key step towards developing countermeasures and therapies for stresses associated with prolonged human exposure to space environments.

Research Impact/Earth Benefits: Along with using mechanisms of stress tolerance to counteract detrimental effects of space travel, data from our proposed experiments could be used in the long term toward solving serious problems in the field of human health. Utilizing mechanisms that allow tardigrades to stabilize their cellular proteins and nucleic acids has been proposed as an option for the dry storage and stabilization of vaccines and other biomaterials (Guo et al., 2000; Wolkers et al., 2001; Puhlev et al., 2001). Because current techniques for vaccine production, distribution, and storage nearly always require a constant cold chain (e.g., -80 and 20 degrees C freezers), these processes are extremely expensive. Some estimates put cold chain costs at around 80% of the total cost of vaccination (Chen et al., 2011). By generating additional stress response datasets, such as response to microgravity, freezing, irradiation, and hypoxia, we will increase our ability and that of other researchers to identify specific mediators of desiccation tolerance, which will then be applied to this and similar problems.

Additionally, a better understanding of mechanisms of stress tolerance could lead to the development of drought and/or freeze tolerant crops.

Guo, N., Puhlev, I., Brown, D. R., Mansbridge, J., & Levine, F. (2000). Trehalose expression confers desiccation tolerance on human cells. Nature biotechnology, 18(2), 168-171.

Wolkers, W. F., Walker, N. J., Tablin, F., & Crowe, J. H. (2001). Human platelets loaded with trehalose survive freeze-drying. Cryobiology, 42(2), 79-87.

Puhlev, I., Guo, N., Brown, D. R., & Levine, F. (2001). Desiccation tolerance in human cells. Cryobiology, 42(3), 207-217.

Task Progress & Bibliography Information FY2017 
Task Progress: While awaiting our flight opportunity, we have continued to carry out our ground-based studies examining the mechanisms and mediators used by tardigrades to survive terrestrial abiotic stresses, such as desiccation, freezing, high temperatures, and irradiation. Using results from transcriptomic sequencing we have identified potential mediators of stress tolerance and are evaluating the function and efficacy of these mediators through a combination of reverse genetic, heterologous expression, and biochemical approaches.

Learning more about how tardigrades survive terrestrial abiotic stresses will help build a platform for comparing and contrasting the mediators and mechanisms they use to protect themselves from the harmful effects of spaceflight.

A manuscript has been submitted and is under review for publication in a peer-reviewed journal: Boothby TC, Tapia H, Brozena A, Piszkiewics S, Smith AE, Giovannini I, Rebbechi L, Pielak GJ, Koshland D, Goldstein B. "Tardigrades Use Intrinsically Disordered Proteins to Survive Desiccation."

Bibliography Type: Description: (Last Updated: 07/24/2020)  Show Cumulative Bibliography Listing
 
Articles in Peer-reviewed Journals Boothby TC, Tenlen JR, Smith FW, Wang JR, Patanella KA, Nishimura EO, Tintori SC, Li Q, Jones CD, Yandell M, Messina DN, Glasscock J, Goldstein B. "Evidence for extensive horizontal gene transfer from the draft genome of a tardigrade." Proc Natl Acad Sci U S A. 2015 Dec 29;112(52):15976-81. Epub 2015 Nov 23. http://dx.doi.org/10.1073/pnas.1510461112 ; PubMed PMID: 26598659; PubMed Central PMCID: PMC4702960 , Dec-2015
Articles in Peer-reviewed Journals Boothby TC, Goldstein B. "Reply to Bemm et al. and Arakawa: Identifying foreign genes in independent Hypsibius dujardini genome assemblies." Proc Natl Acad Sci U S A. 2016 May 31;113(22):E3058-61. http://dx.doi.org/10.1073/pnas.1601149113 ; PubMed PMID: 27173900; PubMed Central PMCID: PMC4896697 , May-2016
Project Title:  Using Water Bears to Identify Biological Countermeasures to Stress During Multigenerational Spaceflight Reduce
Images: icon  Fiscal Year: FY 2016 
Division: Space Biology 
Research Discipline/Element:
Space Biology: Cell & Molecular Biology   | Animal Biology: Invertebrate  
Start Date: 11/01/2014  
End Date: 10/31/2017  
Task Last Updated: 08/03/2015 
Download report in PDF pdf
Principal Investigator/Affiliation:   Boothby, Thomas  Ph.D. / University of Wyoming 
Address:  1000 E. University Ave., Department #3944 
 
Laramie , WY 82071 
Email: Thomas.Boothby@uwyo.edu 
Phone:   
Congressional District:
Web:  
Organization Type: UNIVERSITY 
Organization Name: University of Wyoming 
Joint Agency:  
Comments: NOTE: Previously at University of North Carolina until fall 2019.  
Co-Investigator(s)
Affiliation: 
Goldstein, Bob  Ph.D. University of North Carolina 
Project Information: Grant/Contract No. NNX15AB44G 
Responsible Center: NASA ARC 
Grant Monitor: Taylor, Elizabeth  
Center Contact: 650.604.1783 
elizabeth.taylor-23@nasa.gov 
Solicitation: 2014 Space Biology Flight NNH14ZTT001N 
Grant/Contract No.: NNX15AB44G 
Project Type: FLIGHT 
Flight Program: ISS 
TechPort: No 
No. of Post Docs:
No. of PhD Candidates:  
No. of Master's Candidates:  
No. of Bachelor's Candidates:  
No. of PhD Degrees:
No. of Master's Degrees:  
No. of Bachelor's Degrees:  
Space Biology Element: (1) Cell & Molecular Biology
(2) Animal Biology: Invertebrate
Space Biology Cross-Element Discipline: (1) Reproductive Biology
(2) Developmental Biology
Space Biology Special Category: None
Task Description: For most organisms the stresses associated with spaceflight induce a variety of detrimental effects. To foster a safe and productive long-term human presence in space, therapies and countermeasures to spaceflight-induced stress should be developed. Tardigrades (water bears) are polyextremophiles that have evolved to tolerate multiple extreme environments, which are restrictive to most life. In 2007 tardigrades were shown to survive and reproduce normally during an 11-day low Earth orbit on the Foton-M3 Capsule. We speculate that mechanisms tardigrades have evolved to withstand extreme environments on Earth, may as a side-effect, confer protection against the stresses of spaceflight. This makes tardigrades a uniquely valuable system for studying responses to spaceflight. We have sequenced the genome of the tardigrades Hypsibius dujardini, as well as developed and validated experimental and computational approaches for measuring the effect of different environmental conditions on tardigrade gene expression – allowing us to identify mechanisms used by tardigrades to protect themselves from different stresses. We have also developed a reverse genetic approach, RNA interference, for tardigrades that allows us to directly investigate the role of a gene in conferring tolerance to an environment. We will use these approaches to study tardigrades’ initial, as well as multigenerational response to spaceflight and use RNA interference to test the functionality of the genes identified in our study. Next-generation transcriptome sequencing will be conducted on tardigrades cultures kept 0 generations (founding generation) and 4 generations onboard the International Space Station (ISS). Differential expression analysis will be conducted to compare ISS spaceflight timepoints, ground controls, and tardigrades exposed to other extreme stresses (e.g., desiccation, freezing). This approach will allow us to identify potential mediators of stress tolerance, which will serve as candidates for functional RNA interference experiments. Understanding how tardigrades tolerate spaceflight will better guide future research into countermeasures and therapies for humans exposed to the stresses of prolonged space travel. This proposal's strengths are: the use of an organism that is suited to studying mechanisms of multigenerational tolerance of extreme environments and that has an established RNA interference method for confirming the function of genes identified in our study, our Preliminary Results that validate our proposed approach and technical capabilities as well as the uniqueness and suitability of tardigrades that will allow us to conduct this study. The participants for this study are comprised of experts in tardigrades' stress response and have considerable experience with next-generation sequencing and analysis of non-model organisms. The proposed experiments directly address recommendation AH16 of the Decadal Survey and are in line with recommendation OCB-5 and CMM-5 of NASA’s Multigenerational and Developmental Biology of Invertebrates Research Emphasis as well as NASA’s Fundamental Space Biology Plan 2010-2020 goals. Completion of our proposal will identify genes required for tardigrades to survive multigenerational spaceflight and will be a key step towards developing countermeasures and therapies for stresses associated with prolonged human exposure to space environments.

Research Impact/Earth Benefits: Along with using mechanisms of stress tolerance to counteract detrimental effects of space travel, data from our proposed experiments could be used in the long term toward solving serious problems in the field of human health. Utilizing mechanisms that allow tardigrades to stabilize their cellular proteins and nucleic acids has been proposed as an option for the dry storage and stabilization of vaccines and other biomaterials (Guo et al., 2000; Wolkers et al., 2001; Puhlev et al., 2001). Because current techniques for vaccine production, distribution, and storage nearly always require a constant cold chain (e.g., -80 and 20 0C freezers), these processes are extremely expensive. Some estimates put cold chain costs at around 80% of the total cost of vaccination (Chen et al., 2011). By generating additional stress response datasets, such as response to microgravity, freezing, irradiation, and hypoxia, we will increase our ability and that of other researchers to identify specific mediators of desiccation tolerance, which will then be applied to this and similar problems.

Additionally, a better understanding of mechanisms of stress tolerance could lead to the development of drought and/or freeze tolerant crops.

Guo, N., Puhlev, I., Brown, D. R., Mansbridge, J., & Levine, F. (2000). Trehalose expression confers desiccation tolerance on human cells. Nature biotechnology, 18(2), 168-171.

Wolkers, W. F., Walker, N. J., Tablin, F., & Crowe, J. H. (2001). Human platelets loaded with trehalose survive freeze-drying. Cryobiology, 42(2), 79-87.

Puhlev, I., Guo, N., Brown, D. R., & Levine, F. (2001). Desiccation tolerance in human cells. Cryobiology, 42(3), 207-217.

Task Progress & Bibliography Information FY2016 
Task Progress: We have been working to develop and optimize culture and storage conditions for tardigrades onboard the ISS. Below is a summary of our work to date.

Long-term inactivation of cultures by freezing

To better insure survival, synchronize flight and ground experiments, reduce variability in transport temperatures, and to allow for delays in activation as well as precise temporal activation of tardigrade cultures we tested the potential of using freezing as a long-term method of inactivating our tardigrade cultures.

Reproduction of surviving individuals was assessed and in all cases 100% of surviving specimens were found lay viable clutches of eggs.

Freezing in 2mL syringes – survival and dispensing

Since learning that there have been cracking issues with fibercell cartridges used in the BIOS system during freezing we decided to test if freezing our samples in syringes rather than fibercells would be possible.

Additionally we tested the efficiency of inoculating fibercell cartridges using thawed syringe cultures. Dispensing thawed cultures into fibercells was simple and efficient.

Thawing temperatures

To test if the temperature at which samples frozen in syringes are thawed has an effect on survival we froze subcultures in 2mL syringes for 2 weeks. After 2 weeks syringes were placed at various temperatures and allowed to thaw for 45 minutes. Survival was assessed.

Culturing in fibercells

We tested if BIOS culture system is a viable option for culturing tardigrades, we assessed the ability of fibercell cartridges to support long-term tardigrade cultures.

RNAlater fixation time course at -80 degrees C

We have proposed to fix samples using RNAlater to preserve transcriptional profiles until our samples are returned from the International Space Station (ISS). While RNAlater is a well established reagent used for the long-term stabilization of nucleic acids, it has not been tested long-term on tardigrades. To test this we are looking at the integrity of RNA extracted from samples frozen in RNAlater for prescribed periods of time.

Bibliography Type: Description: (Last Updated: 07/24/2020)  Show Cumulative Bibliography Listing
 
 None in FY 2016
Project Title:  Using Water Bears to Identify Biological Countermeasures to Stress During Multigenerational Spaceflight Reduce
Images: icon  Fiscal Year: FY 2015 
Division: Space Biology 
Research Discipline/Element:
Space Biology: Cell & Molecular Biology   | Animal Biology: Invertebrate  
Start Date: 11/01/2014  
End Date: 10/31/2017  
Task Last Updated: 11/25/2014 
Download report in PDF pdf
Principal Investigator/Affiliation:   Boothby, Thomas  Ph.D. / University of Wyoming 
Address:  1000 E. University Ave., Department #3944 
 
Laramie , WY 82071 
Email: Thomas.Boothby@uwyo.edu 
Phone:   
Congressional District:
Web:  
Organization Type: UNIVERSITY 
Organization Name: University of Wyoming 
Joint Agency:  
Comments: NOTE: Previously at University of North Carolina until fall 2019.  
Co-Investigator(s)
Affiliation: 
Goldstein, Bob  Ph.D. University of North Carolina 
Project Information: Grant/Contract No. NNX15AB44G 
Responsible Center: NASA ARC 
Grant Monitor: Smith, Jeffrey  
Center Contact: 650-604-0880 
jeffrey.d.smith2@nasa.gov 
Solicitation: 2014 Space Biology Flight NNH14ZTT001N 
Grant/Contract No.: NNX15AB44G 
Project Type: FLIGHT 
Flight Program: ISS 
No. of Post Docs:  
No. of PhD Candidates:  
No. of Master's Candidates:  
No. of Bachelor's Candidates:  
No. of PhD Degrees:  
No. of Master's Degrees:  
No. of Bachelor's Degrees:  
Space Biology Element: (1) Cell & Molecular Biology
(2) Animal Biology: Invertebrate
Space Biology Cross-Element Discipline: (1) Reproductive Biology
(2) Developmental Biology
Space Biology Special Category: None
Task Description: For most organisms the stresses associated with spaceflight induce a variety of detrimental effects. To foster a safe and productive long-term human presence in space, therapies and countermeasures to spaceflight-induced stress should be developed. Tardigrades (water bears) are polyextremophiles that have evolved to tolerate multiple extreme environments, which are restrictive to most life. In 2007 tardigrades were shown to survive and reproduce normally during an 11-day low Earth orbit on the Foton-M3 Capsule. We speculate that mechanisms tardigrades have evolved to withstand extreme environments on Earth, may as a side-effect, confer protection against the stresses of spaceflight. This makes tardigrades a uniquely valuable system for studying responses to spaceflight. We have sequenced the genome of the tardigrades Hypsibius dujardini, as well as developed and validated experimental and computational approaches for measuring the effect of different environmental conditions on tardigrade gene expression – allowing us to identify mechanisms used by tardigrades to protect themselves from different stresses. We have also developed a reverse genetic approach, RNA interference, for tardigrades that allows us to directly investigate the role of a gene in conferring tolerance to an environment. We will use these approaches to study tardigrades’ initial, as well as multigenerational response to spaceflight and use RNA interference to test the functionality of the genes identified in our study. Next-generation transcriptome sequencing will be conducted on tardigrades cultures kept 0 generations (founding generation) and 4 generations onboard the International Space Station (ISS). Differential expression analysis will be conducted to compare ISS spaceflight timepoints, ground controls, and tardigrades exposed to other extreme stresses (e.g. desiccation, freezing). This approach will allow us to identify potential mediators of stress tolerance, which will serve as candidates for functional RNA interference experiments. Understanding how tardigrades tolerate spaceflight will better guide future research into countermeasures and therapies for humans exposed to the stresses of prolonged space travel. This proposals strengths are: the use of an organism that is suited to studying mechanisms of multigenerational tolerance of extreme environments and that has an established RNA interference method for confirming the function of genes identified in our study, our Preliminary Results that validate our proposed approach and technical capabilities as well as the uniqueness and suitability of tardigrades that will allow us to conduct this study. The participants for this study are comprised of experts in tardigrades stress response and have considerable experience with next-generation sequencing and analysis of non-model organisms. The proposed experiments directly address recommendation AH16 of the Decadal Survey and are in line with recommendation OCB-5 and CMM-5 of NASA’s Multigenerational and Developmental Biology of Invertebrates Research Emphasis as well as NASA’s Fundamental Space Biology Plan 2010-2020 goals. Completion of our proposal will identify genes required for tardigrades to survive multigenerational spaceflight and will be a key step towards developing countermeasures and therapies for stresses associated with prolonged human exposure to space environments.

Research Impact/Earth Benefits: 0

Task Progress & Bibliography Information FY2015 
Task Progress: New project for FY2015.

Bibliography Type: Description: (Last Updated: 07/24/2020)  Show Cumulative Bibliography Listing
 
 None in FY 2015