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1. The tardigrade protein CAHS D interacts with, but does not retain, water in hydrated and desiccated systems

   https://doi.org/10.1038/s41598-023-37485-3  

   Sanchez-Martinez, Silvia; Ramirez, John F; Meese, Emma K; Childs, Charles A; Boothby, Thomas C (December 2023, Scientific Reports)

   Abstract Tardigrades are a group of microscopic animals renowned for their ability to survive near complete desiccation. A family of proteins, unique to tardigrades, called Cytoplasmic Abundant Heat Soluble (CAHS) proteins are necessary to mediate robust desiccation tolerance in these animals. However, the mechanism(s) by which CAHS proteins help to protect tardigrades during water-loss have not been fully elucidated. Here we use thermogravimetric analysis to empirically test the proposed hypothesis that tardigrade CAHS proteins, due to their propensity to form hydrogels, help to retain water during desiccation. We find that regardless of its gelled state, both in vitro and in vivo, a model CAHS protein (CAHS D) retains no more water than common proteins and control cells in the dry state. However, we find that while CAHS D proteins do not increase the total amount of water retained in a dry system, they interact with the small amount of water that does remain. Our study indicates that desiccation tolerance mediated by CAHS D cannot be simply ascribed to water retention and instead implicates its ability to interact more tightly with residual water as a possible mechanism underlying its protective capacity. These results advance our fundamental understanding of tardigrade desiccation tolerance which could provide potential avenues for new technologies to aid in the storage of dry shelf-stable pharmaceuticals and the generation of stress tolerant crops to ensure food security in the face of global climate change. 

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2. Hsp70 chaperones, Ssa1 and Ssa2, limit poly(A) binding protein aggregation

   https://doi.org/10.1091/mbc.E25-01-0027  

   Buchholz, Hannah E; Martin, Sean A; Dorweiler, Jane E; Radtke, Claire M; Knier, Adam S; Beans, Natalia B; Manogaran, Anita L (June 2025, Molecular Biology of the Cell)

   Thibault, Guillaume (Ed.)

   Molecular chaperones play a central role in maintaining protein homeostasis. The highly conserved Hsp70 family of chaperones have major functions in folding of nascent peptides, protein refolding, and protein aggregate disassembly. In yeast, loss of two Hsp70 proteins, Ssa1 and Ssa2, is associated with decreased cellular growth and shortened lifespan. While heterologous or mutant temperature-sensitive proteins form anomalous large cytoplasmic inclusions in ssa1Δssa2Δ strains, it is unclear how endogenous wild-type proteins behave and are regulated in the presence of limiting Hsp70s. Using the wild-type yeast Poly A binding protein (Pab1), which is involved in mRNA binding and forms stress granules (SGs) upon heat shock, Pab1 forms large inclusions in approximately half of ssa1Δssa2Δ cells in the absence of stress. Overexpression of Ssa1, Hsp104, and Sis1 almost completely limits the formation of these large inclusions in ssa1Δssa2Δ, suggesting that excess Ssa1, Hsp104, and Sis1 can each compensate for the lower levels of Ssa proteins. Upon heat shock, SGs also form in cells whether large Pab1 inclusions are present or not. Surprisingly, cells containing only SGs disassemble faster than wild type, whereas cells with both large inclusions disassemble slower albeit completely. We suspect that disassembly of these large inclusions is linked to the elevated heat shock response and elevated Hsp104 and Sis1 levels in ssa1Δssa2Δ strains. We also observed that wild-type cultures grown to saturation also form large Pab1-GFP inclusions. These inclusions can be partially rescued by overexpression of Ssa1. Taken together, our data suggest that Hsp70 not only plays a role in limiting unwanted protein aggregation in normal cells, but as cells age, the depletion of active Hsp70 possibly underlies the age-related aggregation of endogenous proteins. 

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3. A bacterial expression cloning screen reveals single-stranded DNA-binding proteins as potent desicco-protectants

   https://doi.org/10.1016/j.celrep.2024.114956  

   Hibshman, Jonathan D; Clark-Hachtel, Courtney M; Bloom, Kerry S; Goldstein, Bob (November 2024, Cell Reports)

   Desiccation kills most cells. Some proteins have been identified to help certain cells survive desiccation, but many protein protectants are likely to be unknown. Moreover, the mechanisms ensuring protection of key cellular components are incompletely understood. We devised an expression-cloning approach to discover further protectants. We expressed cDNA libraries from two species of tardigrades in E. coli, and we subjected the bacteria to desiccation to select for survivors. Sequencing the populations of surviving bacteria revealed enrichment of mitochondrial single-stranded DNA-binding proteins (mtSSBs) from both tardigrade species. Expression of mtSSBs in bacteria improved desiccation survival as strongly as the best tardigrade protectants known to date. We found that DNA-binding activity of mtSSBs was necessary and sufficient to improve the desiccation tolerance of bacteria. Although tardigrade mtSSBs were among the strongest protectants we found, single-stranded DNA binding proteins in general offered some protection. These results identify single-stranded DNA-binding proteins as potent desicco-protectants. 

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4. Characterizing Gene Copy Number of Heat Shock Protein Gene Families in the Emerald Rockcod, Trematomus bernacchii

   https://doi.org/10.3390/genes11080867  

   Tercero, Anthony D; Place, Sean P (August 2020, Genes)

   The suborder Notothenioidae is comprised of Antarctic fishes, several of which have lost their ability to rapidly upregulate heat shock proteins in response to thermal stress, instead adopting a pattern of expression resembling constitutive genes. Given the cold-denaturing effect that sub-zero waters have on proteins, evolution in the Southern Ocean has likely selected for increased expression of molecular chaperones. These selective pressures may have also enabled retention of gene duplicates, bolstering quantitative output of cytosolic heat shock proteins (HSPs). Given that newly duplicated genes are under more relaxed selection, it is plausible that gene duplication enabled altered regulation of such highly conserved genes. To test for evidence of gene duplication, copy number of various isoforms within major heat shock gene families were characterized via qPCR and compared between the Antarctic notothen, Trematomus bernacchii, which lost the inducible heat shock response, and the non-Antarctic notothen, Notothenia angustata, which maintains an inducible heat shock response. The results indicate duplication of isoforms within the hsp70 and hsp40 super families have occurred in the genome of T. bernacchii. The findings suggest gene duplications may have been critical in maintaining protein folding efficiency in the sub-zero waters and provided an evolutionary mechanism of alternative regulation of these conserved gene families. 

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5. Protein surface chemistry encodes an adaptive tolerance to desiccation

   https://doi.org/10.1016/j.cels.2025.101407  

   Romero-Pérez, Paulette Sofía; Moran, Haley M; Cordone, David P; Horani, Azeem; Truong, Alexander; Manriquez-Sandoval, Edgar; Ramirez, John F; Martinez, Alec; Gollub, Edith; Hunter, Kara; et al (October 2025, Cell Systems)

   Cellular desiccation—the loss of nearly all water from the cell—is a recurring stress that drives widespread protein dysfunction. To survive, part of the proteome must resume function upon rehydration. Which proteins tolerate desiccation, and the molecular determinants that underlie this tolerance, are largely unknown. Here, we use quantitative mass spectrometry and structural proteomics to show that certain proteins possess an innate capacity to tolerate extreme water loss. Structural analyses point to protein surface chemistry as a key determinant of desiccation tolerance, which we test by showing that rational surface mutants can convert a desiccation-sensitive protein into a tolerant one. We also find that highly tolerant proteins are responsible for the production of small-molecule building blocks, while intolerant proteins are involved in energy-consuming processes such as ribosome biogenesis. We propose that this functional bias enables cells to kickstart their metabolism and promote cell survival following desiccation and rehydration. A record of this paper’s transparent peer review process is included in the supplemental information. 

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