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Title: Biophysical Principles Underlying Assembly, Regulation, and Function of Intracellular Condensates
Authors: Lee, Daniel
Advisors: WingreenBrangwynne, NedClifford SP
Contributors: Quantitative Computational Biology Department
Subjects: Biophysics
Issue Date: 2021
Publisher: Princeton, NJ : Princeton University
Abstract: A major hallmark of biology is intricate organization and structural complexity. One crucial example of this is the nucleus, which houses and protects the genome while coordinating gene expression. The disruption of internal nuclear organization is associated with diseases such as progeria and cancer. The internal components of the nucleus are organized by liquid-liquid phase separation (LLPS), wherein proteins and nucleic acids form liquid-like membraneless organelles known as condensates. While the process of LLPS is well-understood in the context of simple mixtures, the complex, non-equilibrium milieu of the cell presents new challenges. In this thesis, we investigate the role of viscoelastic constraints on the assembly of nuclear condensates. First, we combine theory and simulation to show that crosslinked chromatin, the polymeric material into which DNA is packaged, can mechanically suppress condensates' coalescence and ripening, as well as quantitatively control their number, size, and placement. Then, we utilize an engineered optogenetic system to examine the interplay of condensates with the surrounding viscoelastic chromatin network in living cells, demonstrating that condensate growth dynamics are directly inhibited by the chromatin-dense environment, consistent with theoretical scaling arguments and simulations. We also analyze the cluster-size distributions of both engineered and endogenous condensates experimentally and, in combination with simulations, find that the cluster-size distribution can be directly be inferred from condensate production and coagulation dynamics, providing a quantitative framework to elucidate condensate dynamics in cells. Finally, we interrogate transport processes within condensates, finding that the nucleolus, the condensate responsible for ribosomal synthesis, acts as a complex, viscoelastic fluid whose RNA and protein components experience vastly different transport timescales, and that the DNA component of the nucleolus undergoes nonlinear, time-dependent fluctuations, which we attribute to entanglement with slow-moving ribosomal RNAs. Thus, here we describe multiple biological ramifications of phase separation, both in the context of mesoscale assembly as well as internal dynamics.
Alternate format: The Mudd Manuscript Library retains one bound copy of each dissertation. Search for these copies in the library's main catalog:
Type of Material: Academic dissertations (Ph.D.)
Language: en
Appears in Collections:Quantitative Computational Biology

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