Single-molecule imaging reveals dynamic biphasic partition of RNA-binding proteins in stress granules

Autor(en): Niewidok, Benedikt
Igaev, Maxim
da Graca, Abel Pereira
Strassner, Andre
Lenzen, Christine
Richter, Christian P.
Piehler, Jacob 
Kurre, Rainer 
Brandt, Roland 
Stichwörter: BODIES; Cell Biology; CELLS; COMPLEXES; DISEASE; IDENTIFICATION; LIQUID DROPLETS; LOCALIZATION MICROSCOPY; MESSENGER-RNA; PHASE-TRANSITIONS; TRACKING
Erscheinungsdatum: 2018
Herausgeber: ROCKEFELLER UNIV PRESS
Journal: JOURNAL OF CELL BIOLOGY
Volumen: 217
Ausgabe: 4
Startseite: 1303
Seitenende: 1318
Zusammenfassung: 
Stress granules (SGs) are cytosolic, nonmembranous RNA-protein complexes. In vitro experiments suggested that they are formed by liquid-liquid phase separation; however, their properties in mammalian cells remain unclear. We analyzed the distribution and dynamics of two paradigmatic RNA-binding proteins (RBPs), Ras GTPase-activating protein SH3-domain-binding protein (G3BP1) and insulin-like growth factor II mRNA-binding protein 1 (IMP1), with single-molecule resolution in living neuronal cells. Both RBPs exhibited different exchange kinetics between SGs. Within SGs, single-molecule localization microscopy revealed distributed hotspots of immobilized G3BP1 and IMP1 that reflect the presence of relatively immobile nanometer-sized nanocores. We demonstrate alternating binding in nanocores and anomalous diffusion in the liquid phase with similar characteristics for both RBPs. Reduction of low-complexity regions in G3BP1 resulted in less detectable mobile molecules in the liquid phase without change in binding in nanocores. The data provide direct support for liquid droplet behavior of SGs in living cells and reveal transient binding of RBPs in nanocores. Our study uncovers a surprising disconnect between SG partitioning and internal diffusion and interactions of RBPs.
ISSN: 00219525
DOI: 10.1083/jcb.201709007

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