Biology subjects

Brangwynne, C. P.

Publications and source records attributed to Brangwynne, C. P..

3 recordsLinked to original sources

bioRxiv2018

Mapping local and global liquid-liquid phase behavior in living cells using light-activated multivalent seeds

Recent studies show that liquid-liquid phase separation plays a key role in the assembly of diverse intracellular structures. However, the biophysical principles by which phase separation can be precisely localized within subregions of the cell are still largely unclear, particularly for low-abundance proteins. Here we introduce a biomimetic optogenetic system, \"Corelets\", and utilize its rapid and quantitative tunability to map the first full intracellular phase diagrams, which dictate whether phase separation occurs, and if so by nucleation and growth or spinodal decomposition. Surprisingly, both experiments and simulations show that while intracellular concentrations may be insufficient for global phase separation, sequestering protein ligands to slowly diffusing nucleation centers can move the cell into a different region of the phase diagram, resulting in localized phase separation. This diffusive capture mechanism liberates the cell from the constraints of global protein abundance and is likely exploited to pattern condensates associated with diverse biological processes.

biophysics↗

bioRxiv2018

Protein phase separation provides long-term memory of transient spatial stimuli

Protein/RNA clusters arise frequently in spatially-regulated biological processes, from the asymmetric distribution of P granules and PAR proteins in developing embryos to localized receptor oligomers in migratory cells. This co-occurrence suggests that protein clusters might possess intrinsic properties that make them a useful substrate for spatial regulation. Here, we demonstrate that protein droplets show a robust form of spatial memory, maintaining the spatial pattern of an inhibitor of droplet formation long after it has been removed. Despite this persistence, droplets can be highly dynamic, continuously exchanging monomers with the diffuse phase. We investigate the principles of biophysical spatial memory in three contexts: a computational model of phase separation; a novel optogenetic system where light can drive rapid, localized dissociation of liquid-like protein droplets; and membrane-localized signal transduction from clusters of receptor tyrosine kinases. Our results suggest that the persistent polarization underlying many cellular and developmental processes could arise through a simple biophysical process, without any additional requirement for biochemical positive and negative feedback loops.\n\nHighlightsO_LIWe introduce PixELLs, an optogenetic system for protein droplet disassembly.\nC_LIO_LIModeling and experiments demonstrate long-term memory of local droplet dissociation.\nC_LIO_LIDroplets remember spatial stimuli in nuclei, the cytosol and on cell membranes.\nC_LIO_LIFGFR-optoDroplets convert transient local inputs to persistent cytoskeletal responses.\nC_LI

systems biology↗

bioRxiv2017

Microfluidic Immobilization And Subcellular Imaging Of Developing Caenorhabditis elegans

C. elegans has been an essential model organism in the fields of developmental biology, neuroscience, and aging. However, these areas have been limited by our ability to visualize and track individual C. elegans worms, especially at the subcellular scale, over the course of their lifetime. Here we present a microfluidic device to culture individual C. elegans in parallel throughout post-embryonic development. The device allows for periodic mechanical immobilization of the worm, enabling 3D imaging at subcellular precision. The immobilization is sufficient to enable fluorescence recovery after photobleaching (FRAP) measurements on organelles and other substructures within the same specific cells, throughout larval development, without the use of chemical anesthetics. Using this device, we measure FRAP recovery of two nucleolar proteins in specific intestinal cells within the same worms during larval development. We show that these exhibit different fluorescence recovery as they grow, suggesting differential protein interactions during development. We anticipate that this device will help expand the possible uses of C. elegans as a model organism, enabling its use in addressing fundamental questions at the subcellular scale, including the role of phase transitions in driving spatiotemporal intracellular organization within multicellular organisms.

developmental biology↗