Intrinsically Disordered Tardigrade Proteins Self‐Assemble into Fibrous Gels in Response to Environmental Stress

Abstract

Tardigrades are remarkable for their ability to survive harsh stress conditions as diverse as extreme temperature and desiccation. The molecular mechanisms that confer this unusual resistance to physical stress remain unknown. Recently, tardigrade‐unique intrinsically disordered proteins have been shown to play an essential role in tardigrade anhydrobiosis. Here, we characterize the conformational and physical behaviour of CAHS‐8 from Hypsibius exemplaris. NMR spectroscopy reveals that the protein comprises an extended central helical domain flanked by disordered termini. Upon concentration, the protein is shown to successively form oligomers, long fibres, and finally gels constituted of fibres in a strongly temperature‐dependent manner. The helical domain forms the core of the fibrillar structure, with the disordered termini remaining highly dynamic within the gel. Soluble proteins can be encapsulated within cavities in the gel, maintaining their functional form. The ability to reversibly form fibrous gels may be associated with the enhanced protective properties of these proteins.

Natural and designed proteins inspired by extremotolerant organisms can form condensates and attenuate apoptosis in human cells

“…For those proteins with no visible condensates, the experimental conditions may be insufficient to support phase separation. PARRC CAHS2 exhibited the most dramatic phenotype, which is consistent with recent work describing the gel forming capabilities of CAHS proteins 44-45 (Figure 4E&F). ApoE variants were of interest, based on the puncta observed in the top performing ApoE constructs.…”

Systematic identification of conditionally folded intrinsically disordered regions by AlphaFold2

“…Nuclear magnetic resonance (NMR) spectroscopy is well suited to an ensemble-based structural characterization of IDRs at atomic resolution (Jensen et al 2014; Konrat 2014; Mittag & Forman-Kay 2007). The intrinsic dynamics of IDRs often lead to long-lived NMR signals that can be exploited to collect high-quality data (Malki et al 2021; Sugase et al 2007; Theillet et al 2016). Indeed, a battery of NMR experiments have been applied to probe the conformations of IDRs and residual structure therein (Bertoncini et al 2005; Dyson & Wright 2021; Eliezer 2007; Kakeshpour et al 2021; Mantsyzov et al 2014; Salmon et al 2010), with dedicated software programs focused on integrating NMR and other biophysical methods to determine ensemble representations of IDPs that best agree with the experimental data (Bottaro et al 2020; Choy & Forman-Kay 2001; Gomes et al 2020; Krzeminski et al 2013; Lincoff et al 2020; Ozenne et al 2012; Salmon et al 2010).…”

Desiccation‐tolerance and globular proteins adsorb similar amounts of water

“…These data also suggest that the ability of CAHS D to form hydrogels 25,26 is not explained by hygroscopicity, because CAHS D adsorbs no more water than nongelling proteins (GB1, PvLEA4, etc. ).…”

“…[22][23][24] They also form reversible, concentration-dependent hydrogels. 25,26 We tested CAHS D from the tardigrade Hypsibius exemplaris. The midge Polypedilum vanderplanki survives multiple cycles of complete dehydration and accumulates late embryogenesis abundant (LEA) proteins upon desiccation.…”

“…Cytosolic abundant heat soluble (CAHS) proteins are unique to tardigrades, necessary for their desiccation survival, intrinsically disordered, and protect heterologously expressing cells and enzymes from desiccation damage 22–24 . They also form reversible, concentration‐dependent hydrogels 25,26 . We tested CAHS D from the tardigrade Hypsibius exemplaris .…”