PhD Candiate , University of California, Berkeley
I study molecular mechanisms of how tardigrades (water bears) can survive extremely cold temperature
I study the evolution and mechanisms of cold adaptation in several species of extremophilic microorganisms known as tardigrades (water bears). Although some species of tardigrades have been shown to survive down to ~-250C (colder than any other known eukaryote), little is known about the specific mechanisms that these tardigrades employ to survive such extreme cold. Interestingly, no one has yet published evidence that tardigrades utilize ice-binding (antifreeze and/or ice-nucleating) proteins—which is a commonly used strategy to combat or carefully control the damage of ice formation in a multitude of less cold hardy Antarctic fishes and temperate insects and animals. In my current work, I use a combination of experimental and computational methods to determine what types of proteins or other molecules tardigrades may utilize to survive freezing temperatures. If tardigrades produce any unique proteins, compounds, or utilize other mechanisms to survive extreme temperatures and freezing, we could learn more not only about their evolutionary history, but we could apply such findings to fields of medicine and biology that could benefit from improving cryopreservation methods. I am currently a third year PhD student at UC Berkeley in Dr. Caroline William's lab, and I have been studying the ecology and biology of tardigrades since I was 14 years old—with the help of long-time tardigrade expert Dr. Diane Nelson and Dr. Ralph Schill. Trying to understand the underlying biology of how tardigrades evolved and survive extreme conditions (especially freezing) has been a life-long passion of mine. Before becoming a PhD student, I also taught numerous workshops on the biology of tardigrades to middle school and high school students through MIT's SPLASH program and at summer science programs in South Korea. Likewise, I guided several science fair projects of middle school students on the topic of tardigrades, when I worked as a 7th Grade Science Teacher in New York City—prior to my graduate work. Thanks for this great opportunity to share my passion for tardigrades and research with others!
Abstract: Many limno-terrestrial tardigrades live in unstable habitats where they experience extreme environmental conditions such as drought, heat and subzero temperatures. Although their stress tolerance is often related only to the anhydrobiotic state, tardigrades can also be exposed to great daily temperature fluctuations without dehydration. Survival of subzero temperatures in an active state requires either the ability to tolerate the freezing of body water or mechanisms to decrease the freezing point. Considering freeze tolerance in tardigrades as a general feature, we studied the survival rate of nine tardigrade species originating from polar, temperate and tropical regions by cooling them at rates of 9, 7, 5, 3 and 1 degrees C h(-1) down to -30 degrees C then returning them to room temperature at 10 degrees C h(-1). The resulting moderate survival after fast and slow cooling rates and low survival after intermediate cooling rates may indicate the influence of a physical effect during fast cooling and the possibility that they are able to synthesize cryoprotectants during slow cooling. Differential scanning calorimetry of starved, fed and cold acclimatized individuals showed no intraspecific significant differences in supercooling points and ice formation. Although this might suggest that metabolic and biochemical preparation are non-essential prior to subzero temperature exposure, the increased survival rate with slower cooling rates gives evidence that tardigrades still use some kind of mechanism to protect their cellular structure from freezing injury without influencing the freezing temperature. These results expand our current understanding of freeze tolerance in tardigrades and will lead to a better understanding of their ability to survive subzero temperature conditions.
Pub.: 03 Mar '09, Pinned: 01 Jul '17
Abstract: In tardigrades, tolerance to low temperature is well known and allows them to cope with subzero temperatures in their environment. Although the ability to tolerate freezing body water has been demonstrated in some tardigrades, freeze tolerance of embryonic stages has been little studied, although this has ecological significance. In this study, we evaluated the subzero temperature survival of five different developmental stages of the eutardigrade species Milnesium tardigradum after freezing to -30 degrees C. Embryos were exposed to five different cooling rates between room temperature and -30 degrees C at 1 degrees C/h, 3 degrees C/h, 5 degrees C/h, 7 degrees C/h, and 9 degrees C/h followed by a warming period at 10 degrees C/h. The results showed that the developmental stage and the cooling rate have a significant effect on the hatching rate. Less developed embryonic stages were more sensitive to freezing at higher freezing rates than more developed stages. Differential Scanning Calorimetry (DSC) was used to determine the temperature of crystallization (Tc) in single embryos of the different developmental stages and revealed no differences between the stages. Based on the calorimetric data, we also conclude that the ice nucleation is homogeneous in embryonic stages in tardigrades, as also recently shown for fully developed tardigrades, and not triggered by nucleating agents.
Pub.: 02 Feb '10, Pinned: 01 Jul '17
Abstract: Tardigrades are small, multicellular invertebrates which are able to survive times of unfavourable environmental conditions using their well-known capability to undergo cryptobiosis at any stage of their life cycle. Milnesium tardigradum has become a powerful model system for the analysis of cryptobiosis. While some genetic information is already available for Milnesium tardigradum the proteome is still to be discovered.Here we present to the best of our knowledge the first comprehensive study of Milnesium tardigradum on the protein level. To establish a proteome reference map we developed optimized protocols for protein extraction from tardigrades in the active state and for separation of proteins by high resolution two-dimensional gel electrophoresis. Since only limited sequence information of M. tardigradum on the genome and gene expression level is available to date in public databases we initiated in parallel a tardigrade EST sequencing project to allow for protein identification by electrospray ionization tandem mass spectrometry. 271 out of 606 analyzed protein spots could be identified by searching against the publicly available NCBInr database as well as our newly established tardigrade protein database corresponding to 144 unique proteins. Another 150 spots could be identified in the tardigrade clustered EST database corresponding to 36 unique contigs and ESTs. Proteins with annotated function were further categorized in more detail by their molecular function, biological process and cellular component. For the proteins of unknown function more information could be obtained by performing a protein domain annotation analysis. Our results include proteins like protein member of different heat shock protein families and LEA group 3, which might play important roles in surviving extreme conditions.The proteome reference map of Milnesium tardigradum provides the basis for further studies in order to identify and characterize the biochemical mechanisms of tolerance to extreme desiccation. The optimized proteomics workflow will enable application of sensitive quantification techniques to detect differences in protein expression, which are characteristic of the active and anhydrobiotic states of tardigrades.
Pub.: 13 Mar '10, Pinned: 01 Jul '17
Abstract: Tardigrades have unique stress-adaptations that allow them to survive extremes of cold, heat, radiation and vacuum. To study this, encoded protein clusters and pathways from an ongoing transcriptome study on the tardigrade Milnesium tardigradum were analyzed using bioinformatics tools and compared to expressed sequence tags (ESTs) from Hypsibius dujardini, revealing major pathways involved in resistance against extreme environmental conditions. ESTs are available on the Tardigrade Workbench along with software and databank updates. Our analysis reveals that RNA stability motifs for M. tardigradum are different from typical motifs known from higher animals. M. tardigradum and H. dujardini protein clusters and conserved domains imply metabolic storage pathways for glycogen, glycolipids and specific secondary metabolism as well as stress response pathways (including heat shock proteins, bmh2, and specific repair pathways). Redox-, DNA-, stress- and protein protection pathways complement specific repair capabilities to achieve the strong robustness of M. tardigradum. These pathways are partly conserved in other animals and their manipulation could boost stress adaptation even in human cells. However, the unique combination of resistance and repair pathways make tardigrades and M. tardigradum in particular so highly stress resistant.
Pub.: 09 May '12, Pinned: 01 Jul '17