TY - JOUR
T1 - Composition-dependent thermodynamics of intracellular phase separation
AU - Riback, Joshua A.
AU - Zhu, Lian
AU - Ferrolino, Mylene C.
AU - Tolbert, Michele
AU - Mitrea, Diana M.
AU - Sanders, David W.
AU - Wei, Ming Tzo
AU - Kriwacki, Richard W.
AU - Brangwynne, Clifford P.
N1 - Funding Information:
Acknowledgements We thank members of the Brangwynne laboratory for discussions and comments on this manuscript. This work was supported by the Howard Hughes Medical Institute, the St Jude Collaborative on Membraneless Organelles, and grants from the National Institutes of Health (NIH) 4D Nucleome Program (U01 DA040601) and the Princeton Center for Complex Materials, a Materials Research Science and Engineering Center supported by the National Science Foundation (NSF) (DMR 1420541). L.Z. was supported by an NSF graduate fellowship (DGE-1656466). R.W.K acknowledges support from the NIH (R01 GM115634, R35 GM131891 and P30 CA021765 (to St Jude Children’s Research Hospital)) and ALSAC. M.T. acknowledges support from the NIH (F32 GM131524). Some images were acquired at the St Jude Cell & Tissue Imaging Center, which is supported by St Jude Children’s Research Hospital and the National Cancer Institute (P30 CA021765); we thank V. Frohlich and J. Peters for technical assistance.
Publisher Copyright:
© 2020, The Author(s), under exclusive licence to Springer Nature Limited.
PY - 2020/5/14
Y1 - 2020/5/14
N2 - Intracellular bodies such as nucleoli, Cajal bodies and various signalling assemblies represent membraneless organelles, or condensates, that form via liquid–liquid phase separation (LLPS)1,2. Biomolecular interactions—particularly homotypic interactions mediated by self-associating intrinsically disordered protein regions—are thought to underlie the thermodynamic driving forces for LLPS, forming condensates that can facilitate the assembly and processing of biochemically active complexes, such as ribosomal subunits within the nucleolus. Simplified model systems3–6 have led to the concept that a single fixed saturation concentration is a defining feature of endogenous LLPS7–9, and has been suggested as a mechanism for intracellular concentration buffering2,7,8,10. However, the assumption of a fixed saturation concentration remains largely untested within living cells, in which the richly multicomponent nature of condensates could complicate this simple picture. Here we show that heterotypic multicomponent interactions dominate endogenous LLPS, and give rise to nucleoli and other condensates that do not exhibit a fixed saturation concentration. As the concentration of individual components is varied, their partition coefficients change in a manner that can be used to determine the thermodynamic free energies that underlie LLPS. We find that heterotypic interactions among protein and RNA components stabilize various archetypal intracellular condensates—including the nucleolus, Cajal bodies, stress granules and P-bodies—implying that the composition of condensates is finely tuned by the thermodynamics of the underlying biomolecular interaction network. In the context of RNA-processing condensates such as the nucleolus, this manifests in the selective exclusion of fully assembled ribonucleoprotein complexes, providing a thermodynamic basis for vectorial ribosomal RNA flux out of the nucleolus. This methodology is conceptually straightforward and readily implemented, and can be broadly used to extract thermodynamic parameters from microscopy images. These approaches pave the way for a deeper understanding of the thermodynamics of multicomponent intracellular phase behaviour and its interplay with the nonequilibrium activity that is characteristic of endogenous condensates.
AB - Intracellular bodies such as nucleoli, Cajal bodies and various signalling assemblies represent membraneless organelles, or condensates, that form via liquid–liquid phase separation (LLPS)1,2. Biomolecular interactions—particularly homotypic interactions mediated by self-associating intrinsically disordered protein regions—are thought to underlie the thermodynamic driving forces for LLPS, forming condensates that can facilitate the assembly and processing of biochemically active complexes, such as ribosomal subunits within the nucleolus. Simplified model systems3–6 have led to the concept that a single fixed saturation concentration is a defining feature of endogenous LLPS7–9, and has been suggested as a mechanism for intracellular concentration buffering2,7,8,10. However, the assumption of a fixed saturation concentration remains largely untested within living cells, in which the richly multicomponent nature of condensates could complicate this simple picture. Here we show that heterotypic multicomponent interactions dominate endogenous LLPS, and give rise to nucleoli and other condensates that do not exhibit a fixed saturation concentration. As the concentration of individual components is varied, their partition coefficients change in a manner that can be used to determine the thermodynamic free energies that underlie LLPS. We find that heterotypic interactions among protein and RNA components stabilize various archetypal intracellular condensates—including the nucleolus, Cajal bodies, stress granules and P-bodies—implying that the composition of condensates is finely tuned by the thermodynamics of the underlying biomolecular interaction network. In the context of RNA-processing condensates such as the nucleolus, this manifests in the selective exclusion of fully assembled ribonucleoprotein complexes, providing a thermodynamic basis for vectorial ribosomal RNA flux out of the nucleolus. This methodology is conceptually straightforward and readily implemented, and can be broadly used to extract thermodynamic parameters from microscopy images. These approaches pave the way for a deeper understanding of the thermodynamics of multicomponent intracellular phase behaviour and its interplay with the nonequilibrium activity that is characteristic of endogenous condensates.
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U2 - 10.1038/s41586-020-2256-2
DO - 10.1038/s41586-020-2256-2
M3 - Article
C2 - 32405004
AN - SCOPUS:85084626379
SN - 0028-0836
VL - 581
SP - 209
EP - 214
JO - Nature
JF - Nature
IS - 7807
ER -