Biology subjects

Style, R. W.

Publications and source records attributed to Style, R. W..

3 recordsLinked to original sources

bioRxiv2022

A Phase-separating Thiol-ene Photoresin for Volumetric Bioprinting of Macroporous Hydrogels

Macroporous scaffolds facilitate solute transport and cell-cell communication, but materials allowing for in situ pore formation and 3D printing in aqueous solutions are scarce. Here, we introduce an efficient thiol-ene photoclick resin for light-assisted fabrication of cell-compatible macroporous hydrogels via photopolymerization-induced phase separation (PIPS). This resin consists of norbornene-functionalized polyvinyl alcohol, di-thiol crosslinker and dextran sulfate, which can rapidly form a hydrogel with interconnected pores by PIPS. The pore size is tunable in the range of 2-40 m as a function of light intensity, polymer composition and molecular charge. Unlike conventional methods to porous materials, PIPS uniquely allows in situ pore formation in the presence of living cells, thereby enabling 3D cell culture and bioprinting applications. We demonstrate fast 3D photoencapsulation of living cells, enhanced cell spreading in macroporous hydrogels, and tomographic volumetric bioprinting of cm-scale hydrogel constructs with hierarchical pores within 20 seconds. Collectively, this resin is cell-compatible, low-cost, easy-to-make and highly efficient for PIPS, offering promises for fast photofabrication of living tissues with complex porous structures.

bioengineering↗

bioRxiv2021

Non-specific adhesive forces between filaments and membraneless organelles

Membraneless organelles are liquid-like domains that form inside living cells by phase-separation. While standard physical models of their formation assume their surroundings to be a simple liquid, the cytoplasm is an active viscoelastic environment. To investigate potential coupling of phase separation with the cytoskeleton, we quantify structural correlations of stress granules and microtubules in a human-derived epithelial cell line. We find that microtubule networks are significantly perturbed in the vicinity of stress granules, and that large stress granules conform to the local pore-structure of the microtubule network. When microtubules are depolymerized by nocodazole, tubulin enrichment is localized near the surface of stress granules. We interpret these data using a thermodynamic model of partitioning of particles to the surface and bulk of droplets. This analysis shows that proteins generically have a non-specific affinity for droplet interfaces, which becomes most apparent when they weakly partition to the bulk of droplets and have a large molecular weight. In this framework, our data is consistent with a weak ([lsim] kbT) affinity of tubulin sub-units for stress granule interfaces. As microtubules polymerize their affinity for interfaces increases, providing sufficient adhesion to deform droplets and/or the network. We validate this basic physical phenomena in vitro through the interaction of a simple protein-RNA condensate with tubulin and microtubules.

biophysics↗

bioRxiv2021

Sustained Enzymatic Activity and Flow in Crowded Protein Droplets

Living cells harvest energy from their environments to drive the chemical processes that enable life. We introduce a minimal system that operates at similar protein concentrations, metabolic densities, and length scales as living cells. This approach takes advantage of the tendency of phase-separated protein droplets to strongly partition enzymes, while presenting minimal barriers to transport of small molecules across their interface. By dispersing these microreactors in a reservoir of substrate-loaded buffer, we achieve steady states at metabolic densities that match those of the hungriest microorganisms. We further demonstrate the formation of steady pH gradients, capable of driving microscopic flows. Our approach enables the investigation of the function of diverse enzymes in environments that mimic cytoplasm, and provides a flexible platform for studying the collective behavior of matter driven far from equilibrium.

biophysics↗