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      Protein phase separation in plant membrane biology: more than just a compartmentalization strategy

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      The Plant Cell
      Oxford University Press (OUP)

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          Abstract

          The formation of biomolecular condensates through phase separation is an important strategy to compartmentalize cellular functions. While it is now well established that condensates exist throughout eukaryotic cells, how condensates assemble and function on lipid membranes is only beginning to be understood. In this perspective, we highlight work from plant, animal, and yeast model systems showing that condensates assemble on many endomembrane surfaces to carry out diverse functions. In vesicle trafficking, condensation has reported roles in the formation of endocytic vesicles and autophagosomes and in the inactivation of secretory COPII vesicles. We briefly discuss how membranes and membrane lipids regulate the formation and function of membrane-associated condensates. This includes how membranes act as surfaces for condensate assembly, with lipids mediating the nucleation of condensates during endocytosis and other processes. Additionally, membrane–condensate interactions give rise to the biophysical property of “wetting”, which has functional importance in shaping autophagosomal and vacuolar membranes. We also speculate on the existence of membrane-associated condensates during cell polarity in plants and discuss how condensation may help to establish functional plasma membrane domains. Lastly, we provide advice on relevant in vitro and in vivo approaches and techniques to study membrane-associated phase separation.

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          Biomolecular condensates: organizers of cellular biochemistry

          In addition to membrane-bound organelles, eukaryotic cells feature various membraneless compartments, including the centrosome, the nucleolus and various granules. Many of these compartments form through liquid–liquid phase separation, and the principles, mechanisms and regulation of their assembly as well as their cellular functions are now beginning to emerge.
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            Considerations and Challenges in Studying Liquid-Liquid Phase Separation and Biomolecular Condensates

            Evidence is now mounting that liquid-liquid phase separation (LLPS) underlies the formation of membraneless compartments in cells. This realization has motivated major efforts to delineate the function of such biomolecular condensates in normal cells and their roles in contexts ranging from development to age-related disease. There is great interest in understanding the underlying biophysical principles and the specific properties of biological condensates with the goal of bringing insights into a wide range of biological processes and systems. The explosion of physiological and pathological contexts involving LLPS requires clear standards for their study. Here, we propose guidelines for rigorous experimental characterization of LLPS processes in vitro and in cells, discuss the caveats of common experimental approaches, and point out experimental and theoretical gaps in the field.
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              Phase Transitions in the Assembly of Multi-Valent Signaling Proteins

              Cells are organized on length scales ranging from Angstroms to microns. However, the mechanisms by which Angstrom-scale molecular properties are translated to micron-scale macroscopic properties are not well understood. Here we show that interactions between diverse, synthetic multivalent macromolecules (including multi-domain proteins and RNA) produce sharp, liquid-liquid demixing phase separations, generating micron-sized liquid droplets in aqueous solution. This macroscopic transition corresponds to a molecular transition between small complexes and large, dynamic supramolecular polymers. The concentrations needed for phase transition are directly related to valency of the interacting species. In the case of the actin regulatory protein, neuronal Wiskott-Aldrich Syndrome Protein (N-WASP) interacting with its established biological partners Nck and phosphorylated nephrin 1 , the phase transition corresponds to a sharp increase in activity toward the actin nucleation factor, Arp2/3 complex. The transition is governed by the degree of phosphorylation of nephrin, explaining how this property of the system can be controlled to regulatory effect by kinases. The widespread occurrence of multivalent systems suggests that phase transitions are likely used to spatially organize and biochemically regulate information throughout biology.
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                Author and article information

                Contributors
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                Journal
                The Plant Cell
                Oxford University Press (OUP)
                1040-4651
                1532-298X
                September 2023
                September 01 2023
                June 23 2023
                September 2023
                September 01 2023
                June 23 2023
                : 35
                : 9
                : 3162-3172
                Article
                10.1093/plcell/koad177
                37352127
                87c16c83-b7ab-4f67-a0ce-d19382f236a0
                © 2023

                https://academic.oup.com/pages/standard-publication-reuse-rights

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