期刊
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
卷 112, 期 23, 页码 7189-7194出版社
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1504822112
关键词
liquid droplets; intracellular phase transition; intrinsically disordered proteins; RNA granules
资金
- Human Frontier Science Program [RGP0007/2012]
- National Institutes of Health New Innovator Awards [1DP2GM105437-01, 1DP2 GM105453]
- Searle Scholars Program
- National Science Foundation CAREER Award [1253035]
- American Cancer Society [RSG-12-066-01-DMC]
- Deutsche Forschungsgemeinschaft [EC369-3/1]
- state of Sachsen-Anhalt (Europaischer Fonds fur Regionale Entwicklung start-up grant)
- Division Of Physics
- Direct For Mathematical & Physical Scien [1253035] Funding Source: National Science Foundation
P granules and other RNA/protein bodies are membrane-less organelles that may assemble by intracellular phase separation, similar to the condensation of water vapor into droplets. However, the molecular driving forces and the nature of the condensed phases remain poorly understood. Here, we show that the Caenorhabditis elegans protein LAF-1, a DDX3 RNA helicase found in P granules, phase separates into P granule-like droplets in vitro. We adapt a microrheology technique to precisely measure the viscoelasticity of micrometer-sized LAF-1 droplets, revealing purely viscous properties highly tunable by salt and RNA concentration. RNA decreases viscosity and increases molecular dynamics within the droplet. Single molecule FRET assays suggest that this RNA fluidization results from highly dynamic RNA-protein interactions that emerge close to the droplet phase boundary. We demonstrate than an N-terminal, arginine/glycine rich, intrinsically disordered protein (IDP) domain of LAF-1 is necessary and sufficient for both phase separation and RNA-protein interactions. In vivo, RNAi knockdown of LAF-1 results in the dissolution of P granules in the early embryo, with an apparent submicromolar phase boundary comparable to that measured in vitro. Together, these findings demonstrate that LAF-1 is important for promoting P granule assembly and provide insight into the mechanism by which IDP-driven molecular interactions give rise to liquid phase organelles with tunable properties.
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