Dynamics of Synthetic Membraneless Organelles in Microfluidic Droplets

Linsenmeier, Miriam; Kopp, Martina; Grigolato, Fulvio; Emmanoulidis, Leonidas; Liu, Dany; Zürcher, Dominik; Hondele, Maria; Weis, Karsten; Palmiero, Umberto Capasso; Arosio, Paolo

doi:10.1002/anie.201907278

Cited by 68 publications

(80 citation statements)

References 31 publications

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“…Linsenmeier et al developed a droplet-based microfluidics by introducing a hydrogel matrix to mimic the cytoskeleton for studying the dynamics of synthetic membraneless organelles of artificial cells (shown in Figure 5A). They observed the nucleation, growth, and coarsening in volumes comparable to cells, and they found that the timescale of phase separation decreases linearly as the volume of compartment increases [120].…”

Section: Hydrogel Droplet For Biomedical Cell Encapsulation and Analysismentioning

confidence: 99%

“…The biomedical application of high-throughput biomedical microfluidic systems (HTBMS) with ECM. (A) A droplet-based microfluidic system was developed by introducing a hydrogel matrix mimicking the cytoskeleton to synthetic membraneless organelles, reproduced with permission from[120]. (B) A combinatorial drug screening system was developed based on 3D-cultured cells.…”

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confidence: 99%

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The Fabrication and Application Mechanism of Microfluidic Systems for High Throughput Biomedical Screening: A Review

Song

et al. 2020

Micromachines

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Microfluidic systems have been widely explored based on microfluidic technology, and it has been widely used for biomedical screening. The key parts are the fabrication of the base scaffold, the construction of the matrix environment in the 3D system, and the application mechanism. In recent years, a variety of new materials have emerged, meanwhile, some new technologies have been developed. In this review, we highlight the properties of high throughput and the biomedical application of the microfluidic chip and focus on the recent progress of the fabrication and application mechanism. The emergence of various biocompatible materials has provided more available raw materials for microfluidic chips. The material is not confined to polydimethylsiloxane (PDMS) and the extracellular microenvironment is not limited by a natural matrix. The mechanism is also developed in diverse ways, including its special physical structure and external field effects, such as dielectrophoresis, magnetophoresis, and acoustophoresis. Furthermore, the cell/organ-based microfluidic system provides a new platform for drug screening due to imitating the anatomic and physiologic properties in vivo. Although microfluidic technology is currently mostly in the laboratory stage, it has great potential for commercial applications in the future.

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Section: Hydrogel Droplet For Biomedical Cell Encapsulation and Analysismentioning

confidence: 99%

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confidence: 99%

The Fabrication and Application Mechanism of Microfluidic Systems for High Throughput Biomedical Screening: A Review

Song

et al. 2020

Micromachines

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“…These organelles, which lack membrane boundaries, are formed by liquid–liquid phase separation of proteins and nucleic acids, and are involved in several biological functions and dysfunctions . Most of the proteins associated with these compartments contain low complexity domains (LCDs) or low complexity sequences (LCSs), which are intrinsically disordered domains enriched in specific amino acids . In contrast to complex coacervates, which largely rely on electrostatic interactions, these sequences tune a variety of weak, attractive intermolecular interactions that compete with the entropic cost associated with de‐mixing .…”

Section: Figurementioning

confidence: 99%

Adaptive Chemoenzymatic Microreactors Composed of Inorganic Nanoparticles and Bioinspired Intrinsically Disordered Proteins

et al. 2020

Self Cite

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The assembly of protein and inorganic nanoparticles represents an attractive approach to generate composite materials with multiple functions. Herein, we functionalize inorganic nanoparticles with intrinsically disordered protein domains associated with the formation of membraneless compartments in cells. These protein sequences, defined as low complexity domains (LCDs), encode intermolecular interactions that drive highly controlled, dynamic self‐assembly in response to environmental changes. We show that the properties of the LCDs can be transferred to inorganic nanoparticles, inducing controlled phase separation that is dynamic and responsive to ionic strength and pH. Specifically, we hybridize magnetic nanoparticles with multi‐domain proteins consisting of LCD domains and a globular enzyme, generating dynamic protein‐composite compartments that locally confine hybrid chemoenzymatic reactions and respond to external magnetic fields and changes in solution conditions.

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“…The PhaseScan platform presented herein achieves this objective and allows for the rapid (<5 min), high-throughput, and high-resolution examination of protein LLPS phase behaviour, thereby complementing and expanding upon previous examples of microfluidic investigation of LLPS. 26,29 PhaseScan utilises combinatorial droplet microfluidics to rapidly generate a large number of compartments, each of which can be considered a discrete microenvironment to study protein LLPS. By altering the input solution conditions, a broad range of phaseseparation environments are produced rapidly in order to map LLPS behaviour over a wide region of chemical space.…”

Section: Introductionmentioning

confidence: 99%

High Resolution Biomolecular Condensate Phase Diagrams with a Combinatorial Microdroplet Platform

Arter

Krainer

et al. 2020

Preprint

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The assembly of intracellular proteins into biomolecular condensates via liquid-liquid phase separation (LLPS) has emerged as a fundamental process underlying the organisation and regulation of cellular space and function. Physicochemical characterisation of the parameters that control and modulate phase separation is therefore essential for an improved understanding of protein phase behaviour, including for the therapeutic modulation of LLPS phenomena. A fundamental measure with which to describe protein phase behaviour in chemical space is the phase diagram. Characterisation of phase diagrams requires measuring the presence or absence of the condensed phase under a multitude of conditions and, as such, is associated with significant consumption of time and sample volume even when performed in microwell format.However, due to the rapidly increasing number of biologically and disease-relevant condensate systems, experimental techniques that enable high-throughput analysis of protein phase behaviour are required. To address this challenge, we present here a combinatorial droplet microfluidic platform, termed PhaseScan, for the rapid and high-resolution acquisition of protein phase diagrams. Using this platform, we demonstrate characterisation of the phase behaviour of a pathologically relevant mutant of the protein fused in sarcoma (FUS) in a highly parallelised manner, with significantly improved assay throughput and reduced sample consumption. We demonstrate the capability of the platform by finding the phase boundary at which FUS transitions from a one-phase to a two-phase state as modulated by 1,6-hexanediol, and estimate the free-energy landscape of this system using Flory-Huggins theory. These results thus provide a basis for the rapid acquisition of phase diagrams through the application of microdroplet techniques and pave the way for a wide range of applications, enabling rapid characterisation of the effect of environmental conditions and coacervate species on the thermodynamics of phase separation.

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Dynamics of Synthetic Membraneless Organelles in Microfluidic Droplets

Cited by 68 publications

References 31 publications

The Fabrication and Application Mechanism of Microfluidic Systems for High Throughput Biomedical Screening: A Review

The Fabrication and Application Mechanism of Microfluidic Systems for High Throughput Biomedical Screening: A Review

Adaptive Chemoenzymatic Microreactors Composed of Inorganic Nanoparticles and Bioinspired Intrinsically Disordered Proteins

High Resolution Biomolecular Condensate Phase Diagrams with a Combinatorial Microdroplet Platform

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