Dynamics in the plasma membrane: how to combine fluidity and order
TL;DR: The basic concepts of Brownian diffusion and lipid domain formation in model membranes are summarized and the development of ideas and tools in this field are tracked, outlining key results obtained on the dynamic processes at work in membrane structure and assembly.
Abstract: Cell membranes are fascinating supramolecular aggregates that not only form a barrier between compartments but also harbor many chemical reactions essential to the existence and functioning of a cell. Here, it is proposed to review the molecular dynamics and mosaic organization of the plasma membrane, which are thought to have important functional implications. We will first summarize the basic concepts of Brownian diffusion and lipid domain formation in model membranes and then track the development of ideas and tools in this field, outlining key results obtained on the dynamic processes at work in membrane structure and assembly. We will focus in particular on findings made using fluorescent labeling and imaging procedures to record these dynamic processes. We will also discuss a few examples showing the impact of lateral diffusion on cell signal transduction, and outline some future methodological challenges which must be met before we can answer some of the questions arising in this field of research.
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TL;DR: In this paper, a cyclic disulfide-based functional layer for QCM electrode nanostructuration/functionalization has been shown to provide a robust sensing platform, quite well adapted to the non-planar nature of tested cancer cell monolayers, in a buffered aqueous environment.
Abstract: A Quartz Crystal Microbalance (QCM) is a well-established analytical weighing tool that can serve in the monitoring of the viscoelastic properties of a wide range of chemical and biological analytes, as well as investigating adsorption processes occurring at specifically engineered solid–liquid interfaces. More specifically, several recent works have reported the successful QCM-mediated acoustic profiling of various biophysical transformations of cancerous cells (cancer cell profiling). Such a cancer cell acoustic profiling is aimed at characterizing adsorption, adhesion, spreading, and the homeostatic processes of initially suspended cancer cells, seeded onto various chemically engineered nanostructured QCM sensors. The nanostructuration of sensing the QCM electrode surfaces, with well-defined ligand compositions, provides a unique tool for deeply featuring the physico-chemical interaction basis of multi-parameter bio-molecular recognition events. Even individual recognition ones can be tracked to precisely probe the functional behavior of tested cellular systems. In this regard, the well-recognized 2D organization of self-assembled monolayers (SAMs), of alkane-thiolates/disulfides, onto the bare surfaces of metals, such as gold, generates model chemically defined and structured organic surfaces, for cell interaction, when possessing specific ligands, enabling cell attachment and/or interaction. Herein, and in contrast to the most common suspended cell formats, the real-time in situ sensing, and biochemical profiling, of live raw non-processed cell line monolayers (cell monolayer formats) has been demonstrated to be felicitous. This bio-recognition event used a well-defined chemical composition of a cyclic disulfide-based SAM functional layer for QCM electrode nanostructuration/functionalization. This cyclic disulfide-based functional layer also has been shown to provide a robust sensing platform, quite well adapted to the non-planar nature of tested cancer cell monolayers, in a buffered aqueous environment. Corresponding bio-recognition events, for real-time in situ sensing and profiling, of non-suspended cell monolayers, enabled a significant reduction of the signal processing time, while keeping the interacting cells in their native state. As a matter of consequence, this new QCM-relating development may be considered as a significant step milestone, en-route to multiplexed in/ex vivo real-time diagnostics of tissue pathologies.
8 citations
01 Jan 2017
TL;DR: It is envisioned that unraveling the spatiotemporal complexity of biological membranes would enable us to build a more robust membrane model, which would help in addressing unresolved issues in human health and disease.
Abstract: Biological membranes are complex quasi two-dimensional, supramolecular assemblies of a diverse variety of lipids, proteins and carbohydrates, that compartmentalize living matter into cells and subcellular structures. Membranes are held together by the hydrophobic effect, which is an entropy-driven process originating from strong attractive forces between water molecules. Membrane organization and dynamics are characterized by the absence of intermolecular connectivity among its constituent units, thermodynamically controlled (spontaneous) self assembly, and inherent dynamics characterized by a gradient. Membrane phenomena display a wide range of spatiotemporal scales, thereby making it challenging for experiments and simulations alike. We envision that unraveling the spatiotemporal complexity of biological membranes would enable us to build a more robust membrane model, which would help in addressing unresolved issues in human health and disease.
8 citations
Dissertation•
23 Apr 2015
TL;DR: 2H NMR experiments showed that naphthopyrene can alter membrane properties in the absence of light, and Laurdan general polarization (GP) parameter, which characterizes said spectral shift, has been used to characterize membrane fluidity.
Abstract: The currently accepted model for cell membrane organization involves “lipid rafts”, which differ in composition from the surrounding lipid sea. The existence of these nanoscale heterogeneities is supported by observation of coexisting ordered and disordered lipid phases in lipid model membranes. Fluorescence is a popular family of techniques that can provide dynamic and structural information about membranes. With any probe method, characterization of the effects of fluorescent probe addition on the systems they are used to study is important for the interpretation of experimental data. Comparison can be made between labelled and unlabelled samples using a non-perturbing method, such as deuterium nuclear magnetic resonance spectroscopy (2H NMR). This thesis used 2H NMR to study the effects of an equipartitioning probe, Laurdan, and a non-equipartitioning probe, naphthopyrene, on a well-studied three-component lipid membrane system (35:35:30 dioleoyl-sn-glycero-3-phosphocholine (DOPC)/dipalmitoyl-snglycero-3-phosphocholine-D62 (DPPC-D62)/cholesterol) with a miscibility phase transition. In phase-separated membranes, 0.03-0.6% naphthopyrene disordered lipid chains of DPPCD62 in the liquid-disordered phase, but not of those in the liquid-ordered phase. 0.1-2% Laurdan did not affect the DPPC-D62 in either phase in these membranes. Above the miscibility transition temperature (∼ 22◦C), there is a single homogeneous liquid phase that is not perturbed by the addition of either probe. Laurdan is particularly well suited to the study of phase separation in lipid membranes. It partitions equally well into ordered and disordered lipid phases and displays a polarity-dependent emission spectral shift. Laurdan general polarization (GP) parameter, which characterizes said spectral shift, has been used to characterize membrane fluidity. Two-photon excitation microscopy Laurdan GP images were acquired for membranes with 0.2-2% Laurdan, and Laurdan GP values were found to be strongly correlated with 2H NMR methylene order parameters of DPPC-D62 in the liquidordered phase. Finally, photo-induced phase separation is known to occur in fluorescence microscopy experiments; however, our 2H NMR experiments showed that naphthopyrene can alter membrane properties in the absence of light. The fact that trace amounts of probe (e.g., 0.03-0.6% naphthopyrene) affect lipid molecular order has biological implications: biomolecules present in very small amounts are known to have important functions in cells.
7 citations
Cites background from "Dynamics in the plasma membrane: ho..."
...The existence of lipid rafts in living systems remains contentious [56]....
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TL;DR: In this review, the exciting applications of sub-wavelength apertures towards the sensitive and specific characterisation of molecules are described.
Abstract: Nanometric apertures in a metallic film are easy to produce, robust and highly reproducible nanophotonic devices that possess a number of desirable properties for biophotonics. In this review, we will describe the exciting applications of sub-wavelength apertures towards the sensitive and specific characterisation of molecules.
7 citations
Cites background from "Dynamics in the plasma membrane: ho..."
...Since the diffusion of molecules in the cell plasma membrane is highly sensitive to lateral heterogeneities, monitoring the lateral diffusion of lipids and membrane proteins is highly relevant in order to reveal the structure and the role of membrane heterogeneities (Marguet et al., 2006)....
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...Revealing the dynamic organisation of the cell plasma membrane at the submicron level is a challenging task (Marguet et al., 2006)....
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TL;DR: In this article, a new procedure has been developed which quantitatively analyzes the regionalization of a membrane receptor among the lipid domains of different fluidity in the plasma membrane, which is related to an increase of lipid raft clustering.
7 citations
Cites background from "Dynamics in the plasma membrane: ho..."
...Recent techniques are able to visualize different lipid domains within the plasma membranes of individual cells [8,25,26]....
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References
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TL;DR: Results strongly indicate that the bivalent antibodies produce an aggregation of the surface immunoglobulin molecules in the plane of the membrane, which can occur only if the immunoglOBulin molecules are free to diffuse in the membrane.
Abstract: A fluid mosaic model is presented for the gross organization and structure of the proteins and lipids of biological membranes. The model is consistent with the restrictions imposed by thermodynamics. In this model, the proteins that are integral to the membrane are a heterogeneous set of globular molecules, each arranged in an amphipathic structure, that is, with the ionic and highly polar groups protruding from the membrane into the aqueous phase, and the nonpolar groups largely buried in the hydrophobic interior of the membrane. These globular molecules are partially embedded in a matrix of phospholipid. The bulk of the phospholipid is organized as a discontinuous, fluid bilayer, although a small fraction of the lipid may interact specifically with the membrane proteins. The fluid mosaic structure is therefore formally analogous to a two-dimensional oriented solution of integral proteins (or lipoproteins) in the viscous phospholipid bilayer solvent. Recent experiments with a wide variety of techniqes and several different membrane systems are described, all of which abet consistent with, and add much detail to, the fluid mosaic model. It therefore seems appropriate to suggest possible mechanisms for various membrane functions and membrane-mediated phenomena in the light of the model. As examples, experimentally testable mechanisms are suggested for cell surface changes in malignant transformation, and for cooperative effects exhibited in the interactions of membranes with some specific ligands. Note added in proof: Since this article was written, we have obtained electron microscopic evidence (69) that the concanavalin A binding sites on the membranes of SV40 virus-transformed mouse fibroblasts (3T3 cells) are more clustered than the sites on the membranes of normal cells, as predicted by the hypothesis represented in Fig. 7B. T-here has also appeared a study by Taylor et al. (70) showing the remarkable effects produced on lymphocytes by the addition of antibodies directed to their surface immunoglobulin molecules. The antibodies induce a redistribution and pinocytosis of these surface immunoglobulins, so that within about 30 minutes at 37 degrees C the surface immunoglobulins are completely swept out of the membrane. These effects do not occur, however, if the bivalent antibodies are replaced by their univalent Fab fragments or if the antibody experiments are carried out at 0 degrees C instead of 37 degrees C. These and related results strongly indicate that the bivalent antibodies produce an aggregation of the surface immunoglobulin molecules in the plane of the membrane, which can occur only if the immunoglobulin molecules are free to diffuse in the membrane. This aggregation then appears to trigger off the pinocytosis of the membrane components by some unknown mechanism. Such membrane transformations may be of crucial importance in the induction of an antibody response to an antigen, as well as iv other processes of cell differentiation.
7,790 citations
TL;DR: The new generations of qdots have far-reaching potential for the study of intracellular processes at the single-molecule level, high-resolution cellular imaging, long-term in vivo observation of cell trafficking, tumor targeting, and diagnostics.
Abstract: Research on fluorescent semiconductor nanocrystals (also known as quantum dots or qdots) has evolved over the past two decades from electronic materials science to biological applications. We review current approaches to the synthesis, solubilization, and functionalization of qdots and their applications to cell and animal biology. Recent examples of their experimental use include the observation of diffusion of individual glycine receptors in living neurons and the identification of lymph nodes in live animals by near-infrared emission during surgery. The new generations of qdots have farreaching potential for the study of intracellular processes at the single-molecule level, high-resolution cellular imaging, long-term in vivo observation of cell trafficking, tumor targeting, and diagnostics.
7,499 citations
"Dynamics in the plasma membrane: ho..." refers background in this paper
...The use of fluorescent quantum dots is emerging as a promising alternative to classical fluorescent tags (GFPs and organic fluorophores) (Michalet et al, 2005)....
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...…quantum yields, large molar extinction coefficients, size-dependent tunable emission and high photostability) make them appeal- &2006 European Molecular Biology Organization The EMBO Journal VOL 25 | NO 15 | 2006 3449 ing candidate tags for use with SDT (Dahan et al, 2003; Michalet et al, 2005)....
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TL;DR: This review looks at current methods for preparing QD bioconjugates as well as presenting an overview of applications, and concludes that the potential of QDs in biology has just begun to be realized and new avenues will arise as the ability to manipulate these materials improves.
Abstract: One of the fastest moving and most exciting interfaces of nanotechnology is the use of quantum dots (QDs) in biology. The unique optical properties of QDs make them appealing as in vivo and in vitro fluorophores in a variety of biological investigations, in which traditional fluorescent labels based on organic molecules fall short of providing long-term stability and simultaneous detection of multiple signals. The ability to make QDs water soluble and target them to specific biomolecules has led to promising applications in cellular labelling, deep-tissue imaging, assay labelling and as efficient fluorescence resonance energy transfer donors. Despite recent progress, much work still needs to be done to achieve reproducible and robust surface functionalization and develop flexible bioconjugation techniques. In this review, we look at current methods for preparing QD bioconjugates as well as presenting an overview of applications. The potential of QDs in biology has just begun to be realized and new avenues will arise as our ability to manipulate these materials improves.
5,875 citations
"Dynamics in the plasma membrane: ho..." refers background in this paper
...However, there is still a need to improve the functionalization of QD surfaces, the flexibility for bioconjugations and single irreversible molecular associations between individually tracked molecules (Medintz et al, 2005)....
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Book•
01 Jan 1983
TL;DR: This book is a lucid, straightforward introduction to the concepts and techniques of statistical physics that students of biology, biochemistry, and biophysics must know.
Abstract: This book is a lucid, straightforward introduction to the concepts and techniques of statistical physics that students of biology, biochemistry, and biophysics must know. It provides a sound basis for understanding random motions of molecules, subcellular particles, or cells, or of processes that depend on such motion or are markedly affected by it. Readers do not need to understand thermodynamics in order to acquire a knowledge of the physics involved in diffusion, sedimentation, electrophoresis, chromatography, and cell motility--subjects that become lively and immediate when the author discusses them in terms of random walks of individual particles.
3,041 citations
"Dynamics in the plasma membrane: ho..." refers background in this paper
...Brownian motion is a principle that applies to all biological systems (Berg, 1983): as the result of thermal agitation processes, molecules are constantly on the move, colliding with each other and bouncing back and forth (Figure 1)....
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...…plasma membrane dynamics Brownian motion, diffusion and membrane organization Brownian motion is a principle that applies to all biological systems (Berg, 1983): as the result of thermal agitation processes, molecules are constantly on the move, colliding with each other and bouncing back and…...
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TL;DR: A unified characterization of the best available FPs provides a useful guide in narrowing down the options for biological imaging tools.
Abstract: The recent explosion in the diversity of available fluorescent proteins (FPs) promises a wide variety of new tools for biological imaging. With no unified standard for assessing these tools, however, a researcher is faced with difficult questions. Which FPs are best for general use? Which are the brightest? What additional factors determine which are best for a given experiment? Although in many cases, a trial-and-error approach may still be necessary in determining the answers to these questions, a unified characterization of the best available FPs provides a useful guide in narrowing down the options.
2,933 citations
"Dynamics in the plasma membrane: ho..." refers background in this paper
...As the cDNA encoding the GFP was characterized, a wide variety of monomeric fluorescent proteins have provided attractive potential candidates for monitoring dynamic processes in which different molecular species are simultaneously involved (for a review, see Shaner et al, 2005)....
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