scholarly journals Directed Signaling Cascades in Monodisperse Artificial Eukaryotic Cells

2021 ◽  
Author(s):  
Sunidhi Shetty ◽  
Naresh Yandrapalli ◽  
Kerstin Pinkwart ◽  
Dorothee Krafft ◽  
Tanja Vidaković-Koch ◽  
...  
Keyword(s):  
Hierarchical Structures ◽  
Eukaryotic Cells ◽  
Biochemical Reactions ◽  
Bottom Up ◽  
Membrane Pores ◽  
Enzyme Cascade ◽  
Pdms Channel ◽  
Selective Membrane ◽  
Different Populations ◽  
Minimal Cells

<p>The bottom-up assembly of multi-compartment artificial cells that are able to direct biochemical reactions along a specific spatial pathway remains a considerable engineering challenge. In this work, we address this with a microfluidic platform which is able to produce monodisperse multivesicular vesicles (MVVs) to serve as synthetic eukaryotic cells. Using a two-inlet polydimethylsiloxane (PDMS) channel design to co-encapsulate different populations of liposomes we are able to produce lipid-based MVVs in a high-throughput manner and with three separate inner compartments each containing a different enzyme: α-glucosidase, glucose oxidase, and horseradish peroxidase. We demonstrate the ability of these MVVs to carry out directed chemical communication between the compartments <i>via </i>the reconstitution of size-selective membrane pores. Therefore, the signal transduction, which is triggered externally, follows a specific spatial pathway between the compartments. We use this platform to study the effects of enzyme cascade compartmentalization by direct analytical comparison between bulk, one-, two-, and three-compartment systems. This microfluidic strategy to construct complex hierarchical structures is not only suitable to study compartmentation effects on biochemical reactions but is also applicable for developing advanced drug-delivery systems as well as minimal cells in the field of bottom-up synthetic biology.</p>

scholarly journals Directed Signaling Cascades in Monodisperse Artificial Eukaryotic Cells

2021 ◽  
Author(s):  
Sunidhi Shetty ◽  
Naresh Yandrapalli ◽  
Kerstin Pinkwart ◽  
Dorothee Krafft ◽  
Tanja Vidaković-Koch ◽  
...  
Keyword(s):  
Hierarchical Structures ◽  
Eukaryotic Cells ◽  
Biochemical Reactions ◽  
Bottom Up ◽  
Membrane Pores ◽  
Enzyme Cascade ◽  
Pdms Channel ◽  
Selective Membrane ◽  
Different Populations ◽  
Minimal Cells

<p>The bottom-up assembly of multi-compartment artificial cells that are able to direct biochemical reactions along a specific spatial pathway remains a considerable engineering challenge. In this work, we address this with a microfluidic platform which is able to produce monodisperse multivesicular vesicles (MVVs) to serve as synthetic eukaryotic cells. Using a two-inlet polydimethylsiloxane (PDMS) channel design to co-encapsulate different populations of liposomes we are able to produce lipid-based MVVs in a high-throughput manner and with three separate inner compartments each containing a different enzyme: α-glucosidase, glucose oxidase, and horseradish peroxidase. We demonstrate the ability of these MVVs to carry out directed chemical communication between the compartments <i>via </i>the reconstitution of size-selective membrane pores. Therefore, the signal transduction, which is triggered externally, follows a specific spatial pathway between the compartments. We use this platform to study the effects of enzyme cascade compartmentalization by direct analytical comparison between bulk, one-, two-, and three-compartment systems. This microfluidic strategy to construct complex hierarchical structures is not only suitable to study compartmentation effects on biochemical reactions but is also applicable for developing advanced drug-delivery systems as well as minimal cells in the field of bottom-up synthetic biology.</p>

scholarly journals Microfluidic platform enables tailored translocation and reaction cascades in nanoliter droplet networks

2020 ◽  
Vol 3 (1) ◽  
Author(s):  
Simon Bachler ◽  
Dominik Haidas ◽  
Marion Ort ◽  
Todd A. Duncombe ◽  
Petra S. Dittrich
Keyword(s):  
Synthetic Biology ◽  
Lipid Membranes ◽  
Bottom Up ◽  
Microfluidic Platform ◽  
Alpha Hemolysin ◽  
Droplet Interface Bilayer ◽  
Translocation Experiments ◽  
Fluorescent Molecules ◽  
Reaction Cascades ◽  
Minimal Cells

AbstractIn the field of bottom-up synthetic biology, lipid membranes are the scaffold to create minimal cells and mimic reactions and processes at or across the membrane. In this context, we employ here a versatile microfluidic platform that enables precise positioning of nanoliter droplets with user-specified lipid compositions and in a defined pattern. Adjacent droplets make contact and form a droplet interface bilayer to simulate cellular membranes. Translocation of molecules across membranes are tailored by the addition of alpha-hemolysin to selected droplets. Moreover, we developed a protocol to analyze the translocation of non-fluorescent molecules between droplets with mass spectrometry. Our method is capable of automated formation of one- and two-dimensional droplet networks, which we demonstrated by connecting droplets containing different compound and enzyme solutions to perform translocation experiments and a multistep enzymatic cascade reaction across the droplet network. Our platform opens doors for creating complex artificial systems for bottom-up synthetic biology.

scholarly journals Intracellular Hepadnavirus Nucleocapsids Are Selected for Secretion by Envelope Protein-Independent Membrane Binding

2000 ◽  
Vol 74 (24) ◽  
pp. 11472-11478 ◽  
Author(s):  
Hélène Mabit ◽  
Heinz Schaller
Keyword(s):  
Envelope Protein ◽  
Core Protein ◽  
Membrane Binding ◽  
Dna Viruses ◽  
Virus Particles ◽  
Duck Hepatitis ◽  
Double Stranded Dna ◽  
Selective Membrane ◽  
Membrane Attachment ◽  
Different Populations

ABSTRACT Hepadnaviruses are DNA viruses but, as pararetroviruses, their morphogenesis initiates with the encapsidation of an RNA pregenome, and these viruses have therefore evolved mechanisms to exclude nucleocapsids that contain incompletely matured genomes from participating in budding and secretion. We provide here evidence that binding of hepadnavirus core particles from the cytosol to their target membranes is a distinct step in morphogenesis, discriminating among different populations of intracellular capsids. Using the duck hepatitis B virus (DHBV) and a flotation assay, we found about half of the intracellular capsids to be membrane associated due to an intrinsic membrane-binding affinity. In contrast to free cytosolic capsids, this subpopulation contained largely mature, double-stranded DNA genomes and lacked core protein hyperphosphorylation, both features characteristic for secreted virions. Against expectation, however, the selective membrane attachment observed did not require the presence of the large DHBV envelope protein, which has been considered to be crucial for nucleocapsid-membrane interaction. Furthermore, removal of surface-exposed phosphate residues from nonfloating capsids by itself did not suffice to confer membrane affinity and, finally, hyperphosphorylation was absent from nonenveloped nucleocapsids that were released from DHBV-transfected cells. Collectively, these observations argue for a model in which nucleocapsid maturation, involving the viral genome, capsid structure, and capsid dephosphorylation, leads to the exposure of a membrane-binding signal as a step crucial for selecting the matured nucleocapsid to be incorporated into the capsid-independent budding of virus particles.

scholarly journals Formation of Anion-selective Membrane Pores by Texenomycin A, a Basic Lipopeptaibol Antibiotic.

2002 ◽  
Vol 55 (9) ◽  
pp. 826-828 ◽  
Author(s):  
P. A. GRIGORIEV ◽  
A. BERG ◽  
B. SCHLEGEL ◽  
S. HEINZE ◽  
U. GRÄFE
Keyword(s):  
Membrane Pores ◽  
Selective Membrane

scholarly journals Deep Reinforcement Learning with Hierarchical Structures

2021 ◽  
Author(s):  
Siyuan Li
Keyword(s):  
Reinforcement Learning ◽  
Time Scales ◽  
Hierarchical Structures ◽  
Multiple Time Scales ◽  
Multiple Time ◽  
Top Down ◽  
Bottom Up ◽  
Hierarchical Reinforcement Learning ◽  
Future Work

Hierarchical reinforcement learning (HRL), which enables control at multiple time scales, is a promising paradigm to solve challenging and long-horizon tasks. In this paper, we briefly introduce our work in bottom-up and top-down HRL and outline the directions for future work.

scholarly journals Repulsive electrostatic interactions modulate dense and dilute phase properties of biomolecular condensates

2020 ◽  
Author(s):  
Michael D. Crabtree ◽  
Jack Holland ◽  
Purnima Kompella ◽  
Leon Babl ◽  
Noah Turner ◽  
...  
Keyword(s):  
Germ Cell ◽  
Electrostatic Interactions ◽  
Negative Charge ◽  
Weak Interactions ◽  
Fine Tuning ◽  
Eukaryotic Cells ◽  
Biochemical Reactions ◽  
Net Charge ◽  
Solvent Properties

AbstractLiquid-like membraneless organelles form via multiple, weak interactions between biomolecules. The resulting condensed states constitute novel solvent environments inside eukaryotic cells that partition biomolecules and may favour particular biochemical reactions. Here we demonstrate that, in addition to attractive interactions, repulsive electrostatic interactions modulate condensate properties. We find that net charge modulates the formation, morphology and solvent properties of model Ddx4 condensates in cells and in vitro and that a net negative charge is conserved across germ cell-specific Ddx4 orthologues. This conserved net charge provides a sensitivity to multivalent cations that is not observed in somatic paralogues. The disfavouring effect of a net negative charge in Ddx4 orthologues appears to be offset by increased charge patterning, indicating that fine tuning of both attractive and repulsive interactions can create responsive solvent environments inside biomolecular condensates.

scholarly journals Chemotactic movement of a polarity site enables yeast cells to find their mates

2021 ◽  
Vol 118 (22) ◽  
pp. e2025445118
Author(s):  
Debraj Ghose ◽  
Katherine Jacobs ◽  
Samuel Ramirez ◽  
Timothy Elston ◽  
Daniel Lew
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Computational Model ◽  
Initial Orientation ◽  
Yeast Cells ◽  
Eukaryotic Cells ◽  
Stochastic Effects ◽  
Bottom Up ◽  
Complex Chemical ◽  
Chemical Gradients

How small eukaryotic cells can interpret dynamic, noisy, and spatially complex chemical gradients to orient growth or movement is poorly understood. We address this question using Saccharomyces cerevisiae, where cells orient polarity up pheromone gradients during mating. Initial orientation is often incorrect, but polarity sites then move around the cortex in a search for partners. We find that this movement is biased by local pheromone gradients across the polarity site: that is, movement of the polarity site is chemotactic. A bottom-up computational model recapitulates this biased movement. The model reveals how even though pheromone-bound receptors do not mimic the shape of external pheromone gradients, nonlinear and stochastic effects combine to generate effective gradient tracking. This mechanism for gradient tracking may be applicable to any cell that searches for a target in a complex chemical landscape.

Relationships between hierarchical structure and properties in macromolecular systems

1987 ◽  
Vol 45 ◽  
pp. 440-443
Author(s):  
E. Baer
Keyword(s):  
Uniaxial Tension ◽  
Hierarchical Structure ◽  
Inorganic Material ◽  
Hierarchical Structures ◽  
Bone Collagen ◽  
Structure And Properties ◽  
Connective Tissues ◽  
Scale Level ◽  
Fibrous Protein ◽  
Macromolecular Systems

The most advanced macromolecular materials are found in plants and animals, and certainly the connective tissues in mammals are amongst the most advanced macromolecular composites known to mankind. The efficient use of collagen, a fibrous protein, in the design of both soft and hard connective tissues is worthy of comment. Very crudely, in bone collagen serves as a highly efficient binder for the inorganic hydroxyappatite which stiffens the structure. The interactions between the organic fiber of collagen and the inorganic material seem to occur at the nano (scale) level of organization. Epitatic crystallization of the inorganic phase on the fibers has been reported to give a highly anisotropic, stress responsive, structure. Soft connective tissues also have sophisticated oriented hierarchical structures. The collagen fibers are “glued” together by a highly hydrated gel-like proteoglycan matrix. One of the simplest structures of this type is tendon which functions primarily in uniaxial tension as a reinforced elastomeric cable between muscle and bone.

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