Biomolecular condensates undergo a generic shear-mediated liquid-to-solid transition

Abstract

Membrane-less organelles resulting from liquid–liquid phase separation of biopolymers into intracellular condensates control essential biological functions, including messenger RNA processing, cell signalling and embryogenesis1,2,3,4. It has recently been discovered that several such protein condensates can undergo a further irreversible phase transition, forming solid nanoscale aggregates associated with neurodegenerative disease5,6,7. While the irreversible gelation of protein condensates is generally related to malfunction and disease, one case where the liquid-to-solid transition of protein condensates is functional, however, is that of silk spinning8,9. The formation of silk fibrils is largely driven by shear, yet it is not known what factors control the pathological gelation of functional condensates. Here we demonstrate that four proteins and one peptide system, with no function associated with fibre formation, have a strong propensity to undergo a liquid-to-solid transition when exposed to even low levels of mechanical shear once present in their liquid–liquid phase separated form. Using microfluidics to control the application of shear, we generated fibres from single-protein condensates and characterized their structural and material properties as a function of shear stress. Our results reveal generic backbone–backbone hydrogen bonding constraints as a determining factor in governing this transition. These observations suggest that shear can play an important role in the irreversible liquid-to-solid transition of protein condensates, shed light on the role of physical factors in driving this transition in protein aggregation-related diseases and open a new route towards artificial shear responsive biomaterials.

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Fig. 1: Fibre formation from LLPS proteins and peptides as a result of shear.
Fig. 2: Structural changes in FUS droplets following application of mechanical shear.
Fig. 3: Shear-mediated fibre formation probed by microfluidics.
Fig. 4: Material properties of fibres and proposed model of fibre formation from protein condensates under shear.

Data availability

All relevant data are included in the manuscript and Supplementary Information. More detailed protocols, calculation and analysis are available from the authors upon request.

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Acknowledgements

This work is supported by the Welcome Trust, ERC, Alzheimer Association Zenith, ALS Canada–Brain Canada, Canadian Institutes of Health Research and the Cambridge Centre for Misfolding Diseases. We thank S. Zhang, Y. Lu and K.L. Saar for assistance with the design and fabrication of the microfluidic devices; K.H. Muller for help with flash-freezing and SEM imaging; and A. Alexiadis for discussions regarding fluid mechanics calculations.

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Y.S. and T.P.J.K. conceived and designed the study. Y.S., F.S.R., A.K. and A.L. performed the experiments. Y.S., S.Q., P.S.G.-H., C.I., S.A. and A.K. produced the materials. F.S.R. performed AMF-IR and analysed the data. D.V. simulated the flow field. Y.S. imaged samples under SEM and ran microfluidic experiments and FRAP analysis. A.K. performed tensile strength measurements. Y.S., F.S.R., D.V., A.K. P.S.G.-H., S.A. and T.P.J.K. analysed the data. All authors contributed to the writing of the manuscript.

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Correspondence to Tuomas P. J. Knowles.

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Supplementary Information

Supplementary Figs. 1–5, Table 1, Movie descriptions and refs. 1–5.

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Shen, Y., Ruggeri, F.S., Vigolo, D. et al. Biomolecular condensates undergo a generic shear-mediated liquid-to-solid transition. Nat. Nanotechnol. (2020). https://doi.org/10.1038/s41565-020-0731-4

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