Promiscuous interactions and protein disaggregases determine the material state of stress-inducible RNP granules

Abstract

RNA-protein (RNP) granules have been proposed to assemble by forming solid RNA/protein aggregates or through phase separation into a liquid RNA/protein phase. Which model describes RNP granules in living cells is still unclear. In this study, we analyze P bodies in budding yeast and find that they have liquid-like properties. Surprisingly, yeast stress granules adopt a different material state, which is reminiscent of solid protein aggregates and controlled by protein disaggregases. By using an assay to ectopically nucleate RNP granules, we further establish that RNP granule formation does not depend on amyloid-like aggregation but rather involves many promiscuous interactions. Finally, we show that stress granules have different properties in mammalian cells, where they show liquid-like behavior. Thus, we propose that the material state of RNP granules is flexible and that the solid state of yeast stress granules is an adaptation to extreme environments, made possible by the presence of a powerful disaggregation machine.DOI: http://dx.doi.org/10.7554/eLife.06807.001

Hexanediol: a chemical probe to investigate the material properties of membrane-less compartments

“…Prominent foci of Hsp104-positive aggregates were visible after 1 h. These Hsp104-positive protein aggregates were resistant to hexanediol (Panel C). This is in agreement with the finding that yeast stress granules, which colocalize with Hsp104 and depend on Hsp104 for dissolution [2], are insensitive to hexanediol (Panel C). We next tested whether actin filaments and microtubules are affected by hexanediol treatment (Panel D).…”

“…Hexanediol was shown to perturb FG repeat interactions between nucleoporins in nuclear pores [6] [7] and interactions between RNAbinding proteins in RNA-protein granules (RNP) [2] [8]. These studies suggested that 1,6-hexanediol interferes with weak hydrophobic protein-protein or protein-RNA interactions that are required for these dynamic, liquid-like assemblies to form.…”

“…P bodies are active degradation sites of mRNA and have more liquid-like properties that allow for a fast entry and exit of components from the compartment. In contrast, yeast stress granules are mRNA storage sites, which show more solid-like properties [2]. Understanding the link between the physical properties of membrane-less compartments and their function is of utmost importance, also because transitions into more solid-like states have been linked to age-related diseases [3] [4].…”

Prions, amyloids, and RNA: Pieces of a puzzle

“…Only at subsequent stages these droplets maturate into more stable structures that are more likely to incorporate functional amyloid. [122][123][124] In the framework of this model formation of amyloid might also manifest an alternative, strictly pathogenic pathway. 122,123 Consequently, the lack of negative selection toward the formation of amyloid by wild type or single-mutation proteins is explained either by the functionality of the amyloid state in the granules or exclusively by the need to maintain CBRs / LCRs prone for multivalent interactions.…”

“…120,121 Even though the presence of multiple amyloidogenic proteins with CBRs / LCRs in P-bodies and stress granules led to the hypothesis that functional amyloids are implicated in the biogenesis of these RNPs, 45,111,118,119 so far there is no proof that these proteins are present in RNPs in the amyloid state. 122 Indeed, it appears that initial formation of these RNPs is enthalpy driven, depends on multivalent interactions involving both CBRs/LCRs and RNAbinding domains of proteins and RNAs, and leads to the formation of large RNP complexes in extremely dynamic phase-separated liquidlike droplets. Only at subsequent stages these droplets maturate into more stable structures that are more likely to incorporate functional amyloid.…”