Ideses, Y. ; Brill-Karniely, Y. ; Haviv, L. ; Ben-Shaul, A. ; Bernheim-Groswasser, A. .
Arp2/3 Branched Actin Network Mediates Filopodia-Like Bundles Formation In Vitro.
PLOS ONE 2008,
3.
תקצירDuring cellular migration, regulated actin assembly takes place at the cell leading edge, with continuous disassembly deeper in the cell interior. Actin polymerization at the plasma membrane results in the extension of cellular protrusions in the form of lamellipodia and filopodia. To understand how cells regulate the transformation of lamellipodia into filopodia, and to determine the major factors that control their transition, we studied actin self-assembly in the presence of Arp2/3 complex, WASp-VCA and fascin, the major proteins participating in the assembly of lamellipodia and filopodia. We show that in the early stages of actin polymerization fascin is passive while Arp2/3 mediates the formation of dense and highly branched aster-like networks of actin. Once filaments in the periphery of an aster get long enough, fascin becomes active, linking the filaments into bundles which emanate radially from the aster’s surface, resulting in the formation of star-like structures. We show that the number of bundles nucleated per star, as well as their thickness and length, is controlled by the initial concentration of Arp2/3 complex ([Arp2/3]). Specifically, we tested several values of [Arp2/3] and found that for given initial concentrations of actin and fascin, the number of bundles per star, as well as their length and thickness are larger when [Arp2/3] is lower. Our experimental findings can be interpreted and explained using a theoretical scheme which combines Kinetic Monte Carlo simulations for aster growth, with a simple mechanistic model for bundles’ formation and growth. According to this model, bundles emerge from the aster’s (sparsely branched) surface layer. Bundles begin to form when the bending energy associated with bringing two filaments into contact is compensated by the energetic gain resulting from their fascin linking energy. As time evolves the initially thin and short bundles elongate, thus reducing their bending energy and allowing them to further associate and create thicker bundles, until all actin monomers are consumed. This process is essentially irreversible on the time scale of actin polymerization. Two structural parameters, L, which is proportional to the length of filament tips at the aster periphery and b, the spacing between their origins, dictate the onset of bundling; both depending on [Arp2/3]. Cells may use a similar mechanism to regulate filopodia formation along the cell leading edge. Such a mechanism may allow cells to have control over the localization of filopodia by recruiting specific proteins that regulate filaments length (e. g., Dia2) to specific sites along lamellipodia.
Tzlil, S. ; Murray, D. ; Ben-Shaul, A. .
The ‘‘Electrostatic-Switch’’ Mechanism: Monte Carlo Study Of Marcks-Membrane Interaction.
BIOPHYSICAL JOURNAL 2008,
95, 1745-1757.
תקצירThe binding of the myristoylated alanine-rich C kinase substrate (MARCKS) to mixed, fluid, phospholipid membranes is modeled with a recently developed Monte Carlo simulation scheme. The central domain of MARCKS is both basic (zeta = +13) and hydrophobic (five Phe residues), and is flanked with two long chains, one ending with the myristoylated N-terminus. This natively unfolded protein is modeled as a flexible chain of ‘‘beads’’ representing the amino acid residues. The membranes contain neutral (zeta = 0), monovalent (zeta = -1), and tetravalent (zeta = -4) lipids, all of which are laterally mobile. MARCKS-membrane interaction is modeled by Debye-Huckel electrostatic potentials and semiempirical hydrophobic energies. In agreement with experiment, we find that membrane binding is mediated by electrostatic attraction of the basic domain to acidic lipids and membrane penetration of its hydrophobic moieties. The binding is opposed by configurational entropy losses and electrostatic membrane repulsion of the two long chains, and by lipid demixing upon adsorption. The simulations provide a physical model for how membrane-adsorbed MARCKS attracts several PIP2 lipids (zeta = -4) to its vicinity, and how phosphorylation of the central domain (zeta = +13 to zeta = +7) triggers an ‘‘electrostatic switch’’, which weakens both the membrane interaction and PIP(2) sequestration. This scheme captures the essence of ‘‘discreteness of charge’’ at membrane surfaces and can examine the formation of membrane-mediated multicomponent macromolecular complexes that function in many cellular processes.
Teif, V. B. ; Harries, D. ; Lando, D. Y. ; Ben-Shaul, A. .
Matrix Formalism For Site-Specific Binding Of Unstructured Proteins To Multicomponent Lipid Membranes.
JOURNAL OF PEPTIDE SCIENCE 2008,
14, 368-373.
תקצירWe describe a new approach to calculate the binding of flexible peptides and unfolded proteins to multicomponent lipid membranes. The method is based on the transfer matrix formalism of statistical mechanics recently described as a systematic tool to study DNA-protein-drug binding in gene regulation. Using the energies of interaction of the individual polymer segments with different membrane lipid species and the scaling corrections due to polymer looping, we calculate polymer adsorption characteristics and the degree of sequestration of specific membrane lipids. The method is applied to the effector domain of the MARCKS (myristoylated alanine rich C kinase substrate) protein known to be involved in signal transduction through membrane binding. The calculated binding constants of the MARCKS(151-175) peptide and a series of related peptides to mixed PC/PS/PIP2 membranes are in satisfactory agreement with in vitro experiments. Copyright (c) 2008 European Peptide Society and John Wiley & Sons, Ltd.
Zemel, A. ; Ben-Shaul, A. ; May, S. .
Modulation Of The Spontaneous Curvature And Bending Rigidity Of Lipid Membranes By Interfacially Adsorbed Amphipathic Peptides.
JOURNAL OF PHYSICAL CHEMISTRY B 2008,
112, 6988-6996.
תקצירAmphipathic alpha-helical peptides are often ascribed an ability to induce curvature stress in lipid membranes. This may lead directly to a bending deformation of the host membrane, or it may promote the formation of defects that involve highly curved lipid layers present in membrane pores, fusion intermediates, and solubilized peptide-micelle complexes. The driving force is the same in all cases: peptides induce a spontaneous curvature in the host lipid layer, the sign of which depends sensitively on the peptide’s structural properties. We provide a quantitative account for this observation on the basis of a molecular-level method. To this end, we consider a lipid membrane with peptides interfacially adsorbed onto one leaflet at high peptide-to-lipid ratio. The peptides are modeled generically as rigid cylinders that interact with the host membrane through a perturbation of the conformational properties of the lipid chains. Through the use of a molecular-level chain packing theory, we calculate the elastic properties, that is, the spontaneous curvature and bending stiffness, of the peptide-decorated lipid membrane as a function of the peptide’s insertion depth. We find a positive spontaneous curvature (preferred bending of the membrane away from the peptide) for small penetration depths of the peptide. At a penetration depth roughly equal to half-insertion into the hydrocarbon core, the spontaneous curvature changes sign, implying negative spontaneous curvature (preferred bending of the membrane toward the peptide) for large penetration depths. Despite thinning of the membrane upon peptide insertion, we find an increase in the bending stiffness. We discuss these findings in terms of how the peptide induces elastic stress.
Yoffe, A. M. ; Prinsen, P. ; Gopal, A. ; Knobler, C. M. ; Gelbart, W. M. ; Ben-Shaul, A. .
Predicting The Sizes Of Large Rna Molecules.
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA 2008,
105, 16153-16158.
תקצירWe present a theory of the dependence on sequence of the three-dimensional size of large single-stranded (ss) RNA molecules. The work is motivated by the fact that the genomes of many viruses are large ssRNA molecules-often several thousand nucleotides long-and that these RNAs are spontaneously packaged into small rigid protein shells. We argue that there has been evolutionary pressure for the genome to have overall spatial properties-including an appropriate radius of gyration, R-g-that facilitate this assembly process. For an arbitrary RNA sequence, we introduce the (thermal) average maximum ladder distance (< MLD >) and use it as a measure of the ‘‘extendedness’’ of the RNA secondary structure. The < MLD > values of viral ssRNAs that package into capsids of fixed size are shown to be consistently smaller than those for randomly permuted sequences of the same length and base composition, and also smaller than those of natural ssRNAs that are not under evolutionary pressure to have a compact native form. By mapping these secondary structures onto a linear polymer model and by using < MLD > as a measure of effective contour length, we predict the R-g values of viral ssRNAs are smaller than those of nonviral sequences. More generally, we predict the average < MLD > values of large nonviral ssRNAs scale as N-0.67 +/- 0.01, where N is the number of nucleotides, and that their R-g values vary as < MLD >(0.5) in an ideal solvent, and hence as N-0.34. An alternative analysis, which explicitly includes all branches, is introduced and shown to yield consistent results.