Bulletin of the American Physical Society
APS March Meeting 2023
Volume 68, Number 3
Las Vegas, Nevada (March 5-10)
Virtual (March 20-22); Time Zone: Pacific Time
Session M06: Biomolecular Condesates I - Complex AssembliesFocus
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Sponsoring Units: DBIO Chair: Trevor GrandPre, Princeton University Room: Room 129 |
Wednesday, March 8, 2023 8:00AM - 8:36AM |
M06.00001: Probing the material properties of multi-component and multi-phasic condensates Invited Speaker: Priya R Banerjee The material and dynamical properties of phase-separated biomolecular condensates are key determinants of their biological functions and pathological effects inside living cells. Recent advances indicate that biomolecular condensates are viscoelastic network fluids with time-dependent material properties. Here, we aim to establish a molecular grammar of sequence-encoded and structure-specific protein-protein and protein-RNA interactions that governs the biomolecular condensate phase behavior and dynamical properties. By employing microrheology with optical tweezers, we quantify the viscoelastic dynamics of a variety of binary and ternary biomolecular condensates formed by intrinsically disordered polypeptides (IDPs). We find that at shorter timescales, IDP condensates have an elastically dominant rheological response, while at longer timescales, the same condensates behave as predominantly viscous liquids. The network reconfiguration time, viz., the timescale at which the condensate transitions from an elastically dominant behavior to a viscous behavior, is determined by the IDP sequence composition and patterning, RNA sequence, and RNA secondary structure. Further, by mapping their binary and ternary phase behavior, we show that the viscoelastic behavior and multiphasic dynamics of condensates are collectively determined by the sequence- and structure-encoded biomolecular interactions at the microscopic scale. Overall, our work reveals that condensates are network fluids, and IDP valence and interaction strengths determine the driving forces for phase separation and condensate viscoelastic properties in a predictive manner. |
Wednesday, March 8, 2023 8:36AM - 8:48AM |
M06.00002: Characterizing the structural dynamics of nucleolus during cell cycle Truong An Pham, Madhav Mani, XIAOZHONG Wang, Reza Vafabakhsh Nucleolus is the largest membraneless organelle in the cell. It is the site of rRNA production and ribosome assembly. Nucleolar formation and persistence are governed by the liquid-liquid phase separation paradigm which must persist and adapt through the cell cycle. Here, we used 4D live-cell imaging and histogram-based image analysis to quantify the structural dynamic of nucleolus during the cell cycle. We found that nucleolar disassembly is a multi-step process and is synchronized with the cell cycle through multiple processes. The free energy of protein localization from dense to dilute phase during the disassembly process demonstrates an instability of nucleolus at the onset of mitosis. |
Wednesday, March 8, 2023 8:48AM - 9:00AM Author not Attending |
M06.00003: The effect of cell to cell variablilty in multicomponent protein phase separation Kamal Bhandari Proteins form biomolecular condensates inside cells which contribute to multiple cellular processes and regulatory mechanisms. These condensates, also called membrane-less organelles or cellular puncta, are formed by spontaneous condensation of biomolecules into liquid droplets. Interestingly, most toxic phase separation is the result of multicomponent protein phase separation. Given that concentrations of proteins vary substantially from cell to cell, we investigated how cellular variability affects protein phase separation. To that end, we studied a hypothetical two component protein system in which the primary proteins undergoes phase separation due to weak interaction with an adapter protein. Using Flory-Huggins theory and realistic values of variation in cellular proteins, our model predicts that the cell-to-cell heterogeneity of the adapter protein results the different threshold concentrations for primary proteins to phase separate. We observe that there is no clear cut-off concentration threshold of primary protein for the puncta formation as in single component protein system. Other results are discussed as well. |
Wednesday, March 8, 2023 9:00AM - 9:12AM |
M06.00004: Composition tunes physical properties of multi-component biomolecular condensates Patrick M McCall, Anthony A Hyman, Jan Brugues Biomolecular condensates are membrane-less compartments enriched in proteins and nucleic acids, and are thought to provide unique microenvironments for cellular biochemistry. While design principles are emerging to specify condensate properties through protein sequence, much less is known about how condensate microenvironment can be regulated when the molecular identities are fixed. In multi-component systems, one possible route is through modulation of condensate composition. To explore this, we employ quantitative phase imaging and thermodynamic tie-line measurements to measure the stoichiometry inside model multi-component condensates containing full-length FUS protein and RNA. By systematically tuning the ratio of the total protein and RNA concentrations, we find that stoichiometry inside the condensate is a non-linear function of the total stoichiometry and can be tuned over a wide range. As a probe of the local environment inside the condensate, we measure the fluorescence emission spectra of the solvatochromatic dye Nile Red. We observe significant stoichiometry-dependent shifts in the local emission spectra, suggesting that tuning local composition is sufficient to significantly alter the local microenvironment. We discuss implications for the regulation of biochemical reactions by condensate composition. |
Wednesday, March 8, 2023 9:12AM - 9:24AM |
M06.00005: Catalysis-Induced Phase Separation and Autoregulation of Enzymatic Activity Matthew W Cotton, Ramin Golestanian, Jaime Agudo-Canalejo Liquid-liquid phase separation is an exciting feature of subcellular organisation believed to be vital to the formation of membraneless organelles. It is generally believed that the main drivers of phase separation in such systems are attractive equilibrium interactions which are needed to overcome the entropic costs associated with phase separation. However, we present a thermodynamically consistent model describing the dynamics of a multi-component mixture where one enzyme component catalyzes a reaction between other components. We find that the catalytic activity alone can induce phase separation for sufficiently active systems and large enzymes, without any equilibrium interactions between components. In the limit of fast reaction rates, binodal lines can be calculated using a mapping to an effective free energy. When this catalysis-induced phase separation occurs, the overall catalytic rate in the system is reduced and as such this automatically regulates the enzymatic activity. This points to the biological relevance of this phenomenon and how this effect could be used in synthetic systems. |
Wednesday, March 8, 2023 9:24AM - 9:36AM |
M06.00006: Conformational entropy of intrinsically disordered proteins bars intruders from biomolecular condensates Vladimir Grigorev, Ned S Wingreen, Yaojun Zhang It has recently been discovered that eukaryotic cells are host to a multiplicity of non-membrane-bound compartments, which are termed biomolecular condensates. These phase-separated condensates commonly contain protein components with intrinsically disordered regions (IDRs). While many roles of these IDRs have been proposed and demonstrated in the literature, we suggest here an additional underappreciated role of IDRs, which is to exclude large, unwanted "intruders" from condensates. This exclusion effect arises from the large conformational entropy of IDRs, i.e., there is a large free-energy cost to occupying volume that would otherwise be available to the IDRs. We find that at realistic IDR densities, particles as small as the size of a typical protein (5 nm in diameter) can be more than 90% excluded from condensates. Comparison to data on partitioning of particles into natural and engineered condensates suggests that condensate IDRs may play a generic exclusionary role across organisms and types of condensates. |
Wednesday, March 8, 2023 9:36AM - 9:48AM |
M06.00007: Theory of rheology and aging of protein condensates Ryota Takaki, Marko Popovic, Louise Jawerth, Frank Jülicher The material properties of biological condensates are deemed to play essential roles in cellular functions. The quantitative studies of condensates' rheology, however, became available very recently. The study shows the glass-like behavior of the protein condensates. In this study, we develop a rheological model of biological condensates which shows the experimentally observed features. Starting from physical pictures for the diffusion and stochastic binding of proteins inside condensates, we obtain the constitutive equation for the material property of protein condensates showing aging behavior. We elucidate how aging manifests in the experimental observations in microrheology, both in active and passive rheology. To understand the condensates' aging behavior, we also develop a novel method to characterize the time-dependent rheological properties of aging materials. |
Wednesday, March 8, 2023 9:48AM - 10:00AM |
M06.00008: Material properties of Rubisco-EPYC1 condensates in an algal pyrenoid model Trevor K GrandPre, Yaojun Zhang, Andrew G Pyo, Benjamin Weiner, Ned S Wingreen The photosynthetic alga, Chlamydomonas reinhardtii, concentrates carbon dioxide in the pyrenoid, a biomolecular condensate within the chloroplast, for more efficient energy collection from the sun. The pyrenoid matrix has two main components: the rigid carbon-fixing enzyme Rubisco and a linker protein Essential Pyrenoid Component 1 (EPYC1) which serves as molecular glue. In order to understand the steady-state and dynamical properties of Rubisco-EPYC1 condensates, we developed a coarse-grained model of interacting Rubisco and EPYC1, with bonding kinetics and molecular details learned from experiments. Through this model, we discovered that phase separation depends not only on the bonding stoichiometry between EPYC1 and Rubisco but also on EPYC1 properties such as the number of stickers on each molecule and the linker lengths between stickers. These results help us understand how cells regulate pyrenoid properties such as formation and dissolution via EPYC1 modifications. |
Wednesday, March 8, 2023 10:00AM - 10:12AM |
M06.00009: Engineering biomolecular condensate surfaces Amal Narayanan, Anita Donlic, David W Sanders, Amy R Strom, Ke Xu, Cliff Brangwynne The living cell is a complex and synchronized system with compartmentalization across diverse length scales. These individual compartments coordinate countless coinciding biochemical processes to maintain cell function. Many intracellular organelles such as the lysosome and mitochondria are bound by membranes. But cells also contain organelles that are not confined by membranes, which are known as biomolecular condensates. Over the past decade, growing evidence shows that many biomolecular condensates are viscoelastic materials formed from the phase separation of proteins and nucleic acids. The nucleolus is one condensate that is comprised of multiple immiscible liquid-like layers, which help facilitate the biogenesis of the cell's protein synthesizer – the ribosome. How the interfaces between these distinct sub-phases might control transport and reactivity is important for ribosome biogenesis, and how these interfaces might be modulated to control cancer and other diseases, remains unknown. Here we discuss our work to address this challenge, inspired by native proteins that are specifically enriched at the nucleolar periphery. We show that specific physical and chemical design principles can be exploited to design synthetic proteins that exhibit interfacial localization. These engineered nucleolar surfactants are envisioned as a tool to both elucidate key aspects of the biophysics of condensate interfaces and to enable potential modulation of condensate properties and function. |
Wednesday, March 8, 2023 10:12AM - 10:24AM |
M06.00010: Proximity to criticality determines surface tension of biomolecular condensates Andrew G Pyo, Yaojun Zhang, Ned S Wingreen It has recently become appreciated that cells self-organize their interiors through the formation of biomolecular condensates. These condensates, typically formed through liquid-liquid phase separation of biopolymers, play many functional roles such as signal transduction and selective sequestration. These functions depend on the physical properties of condensates which are encoded in the microscopic features of the constituent biomolecules. Near the critical point, the complex mapping from microscopic features to macroscopic properties can be simplified in terms of a small number of parameters, making it easier to identify underlying principles – but how far does this critical region extend for biomolecular condensates? To address this question, we employed coarse-grained molecular dynamics simulations and found that the critical region is sufficiently large to cover the full physiological range of temperatures. In our model, microscopic features such as polymer sequence strongly affect macroscopic properties, and we were able to trace this sequence dependence simply to a shift in critical temperatures. We also show that the surface tension over a wide range of temperatures can be calculated from the critical temperature and a single measurement of the interface width. |
Wednesday, March 8, 2023 10:24AM - 10:36AM |
M06.00011: Single-molecule trajectories in chemically active condensates Stefano Bo, Frank Jülicher Biomolecular condensates provide distinct chemical environments, which can control various cellular processes. The fluorescent labeling of molecules enables molecular tracking and provides an invaluable tool to probe key processes in cell biology. We discuss how biomolecular condensates govern the kinetics of chemical reactions and how this is reflected in the dynamics of labeled molecules. Our theoretical approach provides insights into how the dynamics of labeled molecules can be used to determine chemical reaction rates inside and outside bimolecular condensates. Finally, we address single-molecule trajectories and relate their statistics to the physics of phase separation. |
Wednesday, March 8, 2023 10:36AM - 10:48AM |
M06.00012: Prewetting of Critical Membranes & Collapsed Polymers Mason N Rouches, Sarah L Veatch, Benjamin B Machta Many intracellular proteins demix into coexisting liquid phases in response to varying cellular conditions. Some, such as those found in growth factor & immune signaling, phase-separate exclusively on the plasma membrane in response to ligand; other nuclear proteins condense exclusively on the 'surface' of chromatin to regulate gene expression. These "surface densities" are typically stable at concentrations far lower than those required without a lower-dimensional template. The surfaces that template these condensates -- chromatin and the plasma membrane -- are poised near phase-transition themselves - in the case of the plasma membrane, near a critical point. Here we propose a minimal theory for phase separation at biological surfaces by cytoplasmic & nuclear proteins. We show that templating surfaces themselves prone to transitions, such as the fluid plasma membrane and dynamically compressible chromatin, significantly expand a regime of "surface-only" phase transitions known as prewetting. We offer two pictures of what a prewetting transition offers to cellular functions. At the plasma membrane we find that prewetting enhances the specificity and sensitivity of signaling networks; in genomic contexts prewetting facilitates precise localization of regulatory machinery |
Wednesday, March 8, 2023 10:48AM - 11:00AM |
M06.00013: Pattern formation of phase-separated lipid domains in membranes Qiwei Yu, Andrej Kosmrlj Giant unilamellar vesicles (GUVs) composed of as few as three lipid species are able to phase separate into small-scale lipid domains on their membranes. Experimentally, the domains are found in both stripes and dots patterns, and the pattern's characteristic size and morphology vary with temperature, membrane tension, and lipid composition [Cornell et al., Biophys. J. 2018]. Here, we present a theoretical model that explains existing observations and makes predictions for future experiments. Our model takes into account the free energy of interactions and mixing of lipids, the elastic deformation energy of membranes, as well as the coupling between the local lipid composition and preferred membrane curvature. This coupling contributes to the selection of a preferred length scale for the emerging phase-separated lipid domains. We also determined the stable morphology of patterns (dots/stripes) by an analysis similar to the Swift-Hohenberg problem. The theory is verified by simulations, which confirm the existence of dots and stripes patterns and elucidate how the resulting patterns depend on parameters that can be controlled in the laboratory. We predict the hysteresis of patterns as a function of these parameters, which can be tested experimentally. Overall, this work helps improve our understanding of pattern formation due to liquid-liquid phase separation on curved surfaces. |
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