Revealing Redox Regulatory Pathways Governing Tardigrade Osmobiosis

Able to survive dehydration through anydrobiosis, freezing temperature through cryobiosis, and extreme osmotic pressure through osmobiosis, tardigrades, otherwise known as waterbears, are valuable organisms to study organismal stress response. However, the signaling pathways involved in their survival are still being uncovered. Herein, we investigate the cysteine-mediated redox regulatory pathways involved in tardigrade osmobiosis induction. We first quantify the reactivity of the tardigrade cysteinome through Quantitative Thiol Reactivity Profiling and reveal for proteins involved in signal transduction pathways to have hyperreactive cysteines. We then quantify cysteine oxidative changes via proteomics during osmobiote formation induced by NaCl and sucrose and uncover that polyamines play a regulatory role during osmobiote formation. An increase in the polyamine spermidine during osmobiosis implicates it as a key player during osmobiosis. Further analysis of the proteins becoming oxidized during osmobiosis reveal that phosphorylation-based signaling from a protein phosphatase 2A-like protein and extracellular signal-regulated kinase 1/2 has a modulatory effect on the induction into and recovery from osmobiosis. In sum, our work is the first to quantify and characterize the tardigrade cysteinome, determine its redox regulation in osmobiosis, and validate its downstream collateral effects during osmobiote formation.