bims-fascar Biomed News
on Phase separation and cellular architecture
Issue of 2020‒11‒22
two papers selected by
Victoria Yan
Max Planck Institute of Molecular Cell Biology and Genetics

  1. Mol Cell. 2020 Nov 09. pii: S1097-2765(20)30736-X. [Epub ahead of print]
      Tethering of synaptic vesicles (SVs) to the active zone determines synaptic strength, although the molecular basis governing SV tethering is elusive. Here, we discover that small unilamellar vesicles (SUVs) and SVs from rat brains coat on the surface of condensed liquid droplets formed by active zone proteins RIM, RIM-BP, and ELKS via phase separation. Remarkably, SUV-coated RIM/RIM-BP condensates are encapsulated by synapsin/SUV condensates, forming two distinct SUV pools reminiscent of the reserve and tethered SV pools that exist in presynaptic boutons. The SUV-coated RIM/RIM-BP condensates can further cluster Ca2+ channels anchored on membranes. Thus, we reconstitute a presynaptic bouton-like structure mimicking the SV-tethered active zone with its one side attached to the presynaptic membrane and the other side connected to the synapsin-clustered SV condensates. The distinct interaction modes between membraneless protein condensates and membrane-based organelles revealed here have general implications in cellular processes, including vesicular formation and trafficking, organelle biogenesis, and autophagy.
    Keywords:  biological condensates; liquid-liquid phase separation; membrane organelles; membraneless condensates; postsynaptic density; presynaptic active zone; synapse formation; synaptic transmission
  2. Cell Cycle. 2020 Nov 18. 1-21
      The centrosome is a unique membraneless organelle that plays a pivotal role in the orderly progression of the cell cycle in animal cells. It has been shown that two pericentriolar scaffold proteins, Cep63 and Cep152, generate a heterotetrameric complex to self-assemble into a higher-order cylindrical architecture around a centriole. However, the mechanisms underlying how they reach their threshold concentrations in the vast intracellular space and generate a self-assembled architecture remain mysterious. Here we demonstrate that, like liquid-like assemblies, Cep63 and Cep152 cooperatively generate amorphous aggregates capable of undergoing dynamic turnover and inter-aggregate fusion in vivo and a significant level of internal rearrangement within a condensate in vitro. Consistently, 1,6-hexanediol, a liquid-liquid phase separation disruptor, greatly diminished the ability of endogenous Cep63 and Cep152 to localize to centrosomes. Interestingly, a purified Cep63•Cep152 complex generated either a cylindrical structure or a vesicle-like hollow sphere in a spatially controlled manner. It also formed condensate-like solid spheres in the presence of a macromolecular crowder. At the molecular level, two hydrophobic motifs, one each from Cep63 and Cep152, were required for generating phase-separating condensates and a high molecular-weight assembly. Thus, we propose that the self-assembly of the Cep63•Cep152 complex is triggered by an intrinsic property of the complex undergoing density transition through the hydrophobic-motif-mediated phase separation. Abbreviations: PCM, pericentriolar material; LLPS, liquid-liquid phase separation; MW, molecular-weight; CLEM, correlative light and electron microscopy; WT, wild-type; CMV, cytomegalovirus; FRAP, fluorescence recovery after photobleaching; FITC, fluorescein isothiocyanate; PCR, polymerase chain reaction; 3D-SIM, three-dimensional structured illumination microscopy; DMEM, Dulbecco's Modified Eagle Medium; PEI Max, Polyethylenimine Max; PBS, phosphate-buffered saline; RT, room temperature; DAPI, 4', 6-diamidino-2-phenylindole; AOTF, acousto-optic tunable filter; LB, Luria broth; OD, optical density; IPTG, isopropyl β-D-1-thiogalactopyranoside; SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
    Keywords:  Cep152; Cep63; PCM; centrosome; phase separation