scholarly journals Actinosomes: condensate-templated proteinaceous containers for engineering synthetic cells

2021 ◽  
Author(s):  
Ketan Ganar ◽  
Liza Leijten ◽  
Siddharth Deshpande
Keyword(s):  
Actin Polymerization ◽  
Three Dimensional ◽  
Active Surface ◽  
Signaling Cascades ◽  
Nucleoside Triphosphate ◽  
Biotechnological Applications ◽  
Effective Technique ◽  
Synthetic Cell ◽  
Synthetic Cells ◽  
Crucial Parameter

Engineering synthetic cells has a broad appeal, from understanding living cells to designing novel biomaterials for therapeutics, biosensing, and hybrid interfaces. A key prerequisite to creating synthetic cells is a functional three-dimensional container capable of orchestrating biochemical reactions. In this study, we present an easy and effective technique to make cell-sized porous containers crafted using the interactions between biomolecular condensates and actin cytoskeleton - we coin them actinosomes. This approach uses polypeptide/nucleoside triphosphate condensates and localizes actin monomers on their surface. By triggering actin polymerization at the expense of sequestered ATP and using osmotic gradients, the condensates are structurally transformed into containers with the boundary made up of actin filaments and polylysine polymers. We show that the GTP-to-ATP ratio is a crucial parameter for forming actinosomes: insufficient ATP prevents condensate dissolution while excess ATP leads to undesired crumpling. The surface of actinosomes lacks any structural order and is porous. We show the functionality of the actinosomes by using them as bioreactors capable of protein synthesis. Actinosomes are a handy addition to the synthetic cell platform, with appealing properties like ease-of-production, inherent encapsulation capacity, and an active surface which holds the potential to trigger signaling cascades and form multicellular assemblies, with potential for medical and biotechnological applications.

scholarly journals On Galerkin Approximations of the Surface Active Quasigeostrophic Equations

2016 ◽  
Vol 46 (1) ◽  
pp. 125-139 ◽  
Author(s):  
Cesar B. Rocha ◽  
William R. Young ◽  
Ian Grooms
Keyword(s):  
Galerkin Approximation ◽  
Three Dimensional ◽  
Active Surface ◽  
Inversion Problem ◽  
Conservation Of Energy ◽  
Galerkin Approximations ◽  
Representation Of Solutions ◽  
Surface Active ◽  
Surface Buoyancy ◽  
Quasigeostrophic Equations

AbstractThis study investigates the representation of solutions of the three-dimensional quasigeostrophic (QG) equations using Galerkin series with standard vertical modes, with particular attention to the incorporation of active surface buoyancy dynamics. This study extends two existing Galerkin approaches (A and B) and develops a new Galerkin approximation (C). Approximation A, due to Flierl, represents the streamfunction as a truncated Galerkin series and defines the potential vorticity (PV) that satisfies the inversion problem exactly. Approximation B, due to Tulloch and Smith, represents the PV as a truncated Galerkin series and calculates the streamfunction that satisfies the inversion problem exactly. Approximation C, the true Galerkin approximation for the QG equations, represents both streamfunction and PV as truncated Galerkin series but does not satisfy the inversion equation exactly. The three approximations are fundamentally different unless the boundaries are isopycnal surfaces. The authors discuss the advantages and limitations of approximations A, B, and C in terms of mathematical rigor and conservation laws and illustrate their relative efficiency by solving linear stability problems with nonzero surface buoyancy. With moderate number of modes, B and C have superior accuracy than A at high wavenumbers. Because B lacks the conservation of energy, this study recommends approximation C for constructing solutions to the surface active QG equations using the Galerkin series with standard vertical modes.

scholarly journals Programmed spatial organization of biomacromolecules into discrete, coacervate-based protocells

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Wiggert J. Altenburg ◽  
N. Amy Yewdall ◽  
Daan F. M. Vervoort ◽  
Marleen H. M. E. van Stevendaal ◽  
Alexander F. Mason ◽  
...  
Keyword(s):  
Spatial Organization ◽  
Nitrilotriacetic Acid ◽  
Binding Motif ◽  
Biologically Relevant ◽  
Enzyme Cascade ◽  
Synthetic Cell ◽  
Synthetic Cells ◽  
Membrane Bound ◽  
High Concentrations ◽  
High Level

AbstractThe cell cytosol is crowded with high concentrations of many different biomacromolecules, which is difficult to mimic in bottom-up synthetic cell research and limits the functionality of existing protocellular platforms. There is thus a clear need for a general, biocompatible, and accessible tool to more accurately emulate this environment. Herein, we describe the development of a discrete, membrane-bound coacervate-based protocellular platform that utilizes the well-known binding motif between Ni2+-nitrilotriacetic acid and His-tagged proteins to exercise a high level of control over the loading of biologically relevant macromolecules. This platform can accrete proteins in a controlled, efficient, and benign manner, culminating in the enhancement of an encapsulated two-enzyme cascade and protease-mediated cargo secretion, highlighting the potency of this methodology. This versatile approach for programmed spatial organization of biologically relevant proteins expands the protocellular toolbox, and paves the way for the development of the next generation of complex yet well-regulated synthetic cells.

scholarly journals Active Surface with Dynamic Microstructures and Hierarchical Gradient Enabled by in situ Pneumatic Control

Micromachines ◽  
2020 ◽  
Vol 11 (11) ◽  
pp. 992
Author(s):  
Jian-Nan Wang ◽  
Benfeng Bai ◽  
Qi-Dai Chen ◽  
Hong-Bo Sun
Keyword(s):  
Large Amplitude ◽  
Three Dimensional ◽  
Active Surface ◽  
Bonding Process ◽  
Dynamic Topography ◽  
Large Area ◽  
Pneumatic Control ◽  
Design Flexibility ◽  
Surface Microstructures

An active surface with an on-demand tunable topography holds great potential for various applications, such as reconfigurable metasurfaces, adaptive microlenses, soft robots and four-dimensional (4D) printing. Despite extensive progress, to achieve refined control of microscale surface structures with large-amplitude deformation remains a challenge. Moreover, driven by the demand of constructing a large area of microstructures with increased complexity—for instance, biomimetic functional textures bearing a three-dimensional (3D) gradient—novel strategies are highly desired. Here, we develop an active surface with a dynamic topography and three-tier height gradient via a strain-tunable mismatching-bonding process. Pneumatic actuation allows for rapid, reversible and uniform regulation of surface microstructures at the centimeter scale. The in-situ modulation facilitates large-amplitude deformation with a maximum tuning range of 185 μm. Moreover, the structural gradient can be modulated by programming the strain value of the bonding process. With our strategy, another two types of surfaces with a four-tier gradient and without gradient were also prepared. By providing active modulation and design flexibility of complicated microstructures, the proposed strategy would unlock more opportunities for a wealth of novel utilizations.

scholarly journals Actin polymerization controls the activation of multidrug efflux at fertilization by translocation and fine-scale positioning of ABCB1 on microvilli

2012 ◽  
Vol 23 (18) ◽  
pp. 3663-3672 ◽  
Author(s):  
Kristen Whalen ◽  
Adam M. Reitzel ◽  
Amro Hamdoun
Keyword(s):  
Cell Surface ◽  
Actin Polymerization ◽  
Sea Urchins ◽  
Three Dimensional ◽  
Cortical Reorganization ◽  
Fine Scale ◽  
Multidrug Efflux ◽  
Cortical Actin ◽  
Contrast Enhanced ◽  
And Function

Fertilization changes the structure and function of the cell surface. In sea urchins, these changes include polymerization of cortical actin and a coincident, switch-like increase in the activity of the multidrug efflux transporter ABCB1a. However, it is not clear how cortical reorganization leads to changes in membrane transport physiology. In this study, we used three-dimensional superresolution fluorescence microscopy to resolve the fine-scale movements of the transporter along polymerizing actin filaments, and we show that efflux activity is established after ABCB1a translocates to the tips of the microvilli. Inhibition of actin poly­merization or bundle formation prevents tip localization, resulting in the patching of ABCB1a at the cell surface and decreased efflux activity. In contrast, enhanced actin polymerization promotes tip localization. Finally, interference with Rab11, a regulator of apical recycling, inhibits activation of efflux activity in embryos. Together our results show that actin-mediated, short-range traffic and positioning of transporters at the cell surface regulates multidrug efflux activity and highlight the multifaceted roles of microvilli in the spatial distribution of membrane proteins.

scholarly journals Synthetic Cell as a Platform for Understanding Membrane-Membrane Interactions

Membranes ◽  
2021 ◽  
Vol 11 (12) ◽  
pp. 912
Author(s):  
Bineet Sharma ◽  
Hossein Moghimianavval ◽  
Sung-Won Hwang ◽  
Allen P. Liu
Keyword(s):  
Membrane Fusion ◽  
Intercellular Communication ◽  
Lipid Vesicles ◽  
Bilayer Membrane ◽  
Future Research ◽  
Membrane Interactions ◽  
Protein Insertion ◽  
Synthetic Cell ◽  
Challenges And Opportunities ◽  
Synthetic Cells

In the pursuit of understanding life, model membranes made of phospholipids were envisaged decades ago as a platform for the bottom-up study of biological processes. Micron-sized lipid vesicles have gained great acceptance as their bilayer membrane resembles the natural cell membrane. Important biological events involving membranes, such as membrane protein insertion, membrane fusion, and intercellular communication, will be highlighted in this review with recent research updates. We will first review different lipid bilayer platforms used for incorporation of integral membrane proteins and challenges associated with their functional reconstitution. We next discuss different methods for reconstitution of membrane fusion and compare their fusion efficiency. Lastly, we will highlight the importance and challenges of intercellular communication between synthetic cells and synthetic cells-to-natural cells. We will summarize the review by highlighting the challenges and opportunities associated with studying membrane–membrane interactions and possible future research directions.

scholarly journals Crotoxin Inhibits Endothelial Cell Functions in Two- and Three-dimensional Tumor Microenvironment

2021 ◽  
Vol 12 ◽  
Author(s):  
Ellen Emi Kato ◽  
Luciana Araújo Pimenta ◽  
Maíra Estanislau Soares de Almeida ◽  
Vanessa Olzon Zambelli ◽  
Marinilce Fagundes dos Santos ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Endothelial Cell ◽  
Actin Polymerization ◽  
Type I Collagen ◽  
Three Dimensional ◽  
Endothelial Cell Migration ◽  
Endothelial Cell Proliferation ◽  
Type I ◽  
Cell Functions ◽  
Extracellular Matrix Components

Antitumor property of Crotoxin (CTX), the major toxin from Crotalus durissus terrificus snake venom, has been demonstrated in experimental animal models and clinical trials. However, the direct action of this toxin on the significant events involved in neovascularization, which are essential for tumor growth and survival, has not been confirmed. This study investigated the effects of CTX on the key parameters of neovascularization in two- and three-dimensional culture models. Murine endothelial cell lines derived from thymus hemangioma (t.End.1) were treated at different concentrations of CTX (6.25–200 nM). Endothelial cell proliferation, cell adhesion, and actin cytoskeletal dynamics on laminin (10 µg/ml), type I collagen (10 µg/ml), and fibronectin (3 µg/ml) were evaluated along with the endothelial cell migration and formation of capillary-like tubes in 3D Matrigel. CTX concentration of 50 nM inhibited tube formation on 3D Matrigel and impaired cell adhesion, proliferation, and migration under both culture medium and tumor-conditioned medium. These actions were not accountable for the loss of cell viability. Inhibition of cell adhesion to different extracellular matrix components was related to the reduction of αv and α2 integrin distribution and cytoskeletal actin polymerization (F-actin), accompanied by inhibition of focal adhesion kinase (FAK), Rac1 (GTPase) signaling proteins, and actin-related protein 2/3 (Arp 2/3) complex. This study proved that CTX inhibits the major events involved in angiogenesis, particularly against tumor stimuli, highlighting the importance of the anti-angiogenic action of CTX in inhibition of tumor progression.

scholarly journals Fully 3D Active Surface with Machine Learning for PET Image Segmentation

2020 ◽  
Vol 6 (11) ◽  
pp. 113 ◽  
Author(s):  
Albert Comelli
Keyword(s):  
Machine Learning ◽  
Head And Neck ◽  
Active Contour ◽  
Mathematical Formulation ◽  
Three Dimensional ◽  
Reconstruction Algorithm ◽  
Active Surface ◽  
Dice Similarity Coefficient ◽  
Volume Reconstruction ◽  
Surface Evolution

In order to tackle three-dimensional tumor volume reconstruction from Positron Emission Tomography (PET) images, most of the existing algorithms rely on the segmentation of independent PET slices. To exploit cross-slice information, typically overlooked in these 2D implementations, I present an algorithm capable of achieving the volume reconstruction directly in 3D, by leveraging an active surface algorithm. The evolution of such surface performs the segmentation of the whole stack of slices simultaneously and can handle changes in topology. Furthermore, no artificial stop condition is required, as the active surface will naturally converge to a stable topology. In addition, I include a machine learning component to enhance the accuracy of the segmentation process. The latter consists of a forcing term based on classification results from a discriminant analysis algorithm, which is included directly in the mathematical formulation of the energy function driving surface evolution. It is worth noting that the training of such a component requires minimal data compared to more involved deep learning methods. Only eight patients (i.e., two lung, four head and neck, and two brain cancers) were used for training and testing the machine learning component, while fifty patients (i.e., 10 lung, 25 head and neck, and 15 brain cancers) were used to test the full 3D reconstruction algorithm. Performance evaluation is based on the same dataset of patients discussed in my previous work, where the segmentation was performed using the 2D active contour. The results confirm that the active surface algorithm is superior to the active contour algorithm, outperforming the earlier approach on all the investigated anatomical districts with a dice similarity coefficient of 90.47 ± 2.36% for lung cancer, 88.30 ± 2.89% for head and neck cancer, and 90.29 ± 2.52% for brain cancer. Based on the reported results, it can be claimed that the migration into a 3D system yielded a practical benefit justifying the effort to rewrite an existing 2D system for PET imaging segmentation.

scholarly journals Targeting the mDia Formin-Assembled Cytoskeleton Is an Effective Anti-Invasion Strategy in Adult High-Grade Glioma Patient-Derived Neurospheres

Cancers ◽  
2019 ◽  
Vol 11 (3) ◽  
pp. 392 ◽  
Author(s):  
Krista Pettee ◽  
Kathryn Becker ◽  
Arthur Alberts ◽  
Kevin Reinard ◽  
Jason Schroeder ◽  
...  
Keyword(s):  
Actin Polymerization ◽  
Rho Gtpase ◽  
Three Dimensional ◽  
High Grade Glioma ◽  
High Grade ◽  
Glioma Patient ◽  
Glioma Invasion ◽  
Who Grade ◽  
Rat Brain Slice

High-grade glioma (HGG, WHO Grade III–IV) accounts for the majority of adult primary malignant brain tumors. Failure of current therapies to target invasive glioma cells partly explains the minimal survival advantages: invasive tumors lack easily-defined surgical margins, and are inherently more chemo- and radioresistant. Much work centers upon Rho GTPase-mediated glioma invasion, yet downstream Rho effector roles are poorly understood and represent potential therapeutic targets. The roles for the mammalian Diaphanous (mDia)-related formin family of Rho effectors have emerged in invasive/metastatic disease. mDias assemble linear F-actin to promote protrusive cytoskeletal structures underlying tumor cell invasion. Small molecule mDia intramimic (IMM) agonists induced mDia functional activities including F-actin polymerization. mDia agonism inhibited polarized migration in Glioblastoma (WHO Grade IV) cells in three-dimensional (3D) in vitro and rat brain slice models. Here, we evaluate whether clinically-relevant high-grade glioma patient-derived neuro-sphere invasion is sensitive to formin agonism. Surgical HGG samples were dissociated, briefly grown as monolayers, and spontaneously formed non-adherent neuro-spheres. IMM treatment dramatically inhibited HGG patient neuro-sphere invasion, both at neuro-sphere embedding and mid-invasion assay, inducing an amoeboid morphology in neuro-sphere edge cells, while inhibiting actin- and tubulin-enriched tumor microtube formation. Thus, mDia agonism effectively disrupts multiple aspects of patient-derived HGG neuro-sphere invasion.

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