Anhydrobiotes are rare microbes, plants and animals that tolerate severe water loss. Understanding the molecular basis for their desiccation tolerance may provide novel insights into stress biology and critical tools for engineering drought-tolerant crops. Using the anhydrobiote, budding yeast, we show that trehalose and Hsp12, a small intrinsically disordered protein (sIDP) of the hydrophilin family, synergize to mitigate completely the inviability caused by the lethal stresses of desiccation. We show that these two molecules help to stabilize the activity and prevent aggregation of model proteins both in vivo and in vitro. We also identify a novel in vitro role for Hsp12 as a membrane remodeler, a protective feature not shared by another yeast hydrophilin, suggesting that sIDPs have distinct biological functions.
All data generated or analysed during this study are included in the manuscript and supporting files.
- Skylar Xantus Kim
- Hugo Tapia
- Gamze Çamdere
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
- Anthony A Hyman, Max Planck Institute of Molecular Cell Biology and Genetics, Germany
© 2018, Kim et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Cyclic GMP-dependent protein kinases (PKGs) are key mediators of the nitric oxide/cGMP signaling pathway that regulates biological functions as diverse as smooth muscle contraction, cardiac function, and axon guidance. Understanding how cGMP differentially triggers mammalian PKG isoforms could lead to new therapeutics that inhibit or activate PKGs, complementing drugs that target nitric oxide synthases and cyclic nucleotide phosphodiesterases in this signaling axis. Alternate splicing of PRKG1 transcripts confers distinct leucine zippers, linkers, and auto-inhibitory pseudo-substrate sequences to PKG Iα and Iβ that result in isoform-specific activation properties, but the mechanism of enzyme auto-inhibition and its alleviation by cGMP is not well understood. Here we present a crystal structure of PKG Iβ in which the auto-inhibitory sequence and the cyclic nucleotide binding domains are bound to the catalytic domain, providing a snapshot of the auto-inhibited state. Specific contacts between the PKG Iβ auto-inhibitory sequence and the enzyme active site help explain isoform-specific activation constants and the effects of phosphorylation in the linker. We also present a crystal structure of a PKG I cyclic nucleotide binding domain with an activating mutation linked to Thoracic Aortic Aneurysms and Dissections. Similarity of this structure to wild type cGMP-bound domains and differences with the auto-inhibited enzyme provide a mechanistic basis for constitutive activation. We show that PKG Iβ auto-inhibition is mediated by contacts within each monomer of the native full-length dimeric protein, and using the available structural and biochemical data we develop a model for the regulation and cooperative activation of PKGs.
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