Biomedical subjects

William M Gelbart

Publications and source records attributed to William M Gelbart.

At least 19 recordsLinked to original sources

PubMed2006

VectorBase: a home for invertebrate vectors of human pathogens.

VectorBase (http://www.vectorbase.org/) is a web-accessible data repository for information about invertebrate vectors of human pathogens. VectorBase annotates and maintains vector genomes providing an integrated resource for the research community. Currently, VectorBase contains genome information for two organisms: Anopheles gambiae, a vector for the Plasmodium protozoan agent causing malaria, and Aedes aegypti, a vector for the flaviviral agents causing Yellow fever and Dengue fever.

Aedes↗

PubMed2006

FlyBase: genomes by the dozen.

FlyBase (http://flybase.org/) is the primary database of genetic and genomic data for the insect family Drosophilidae. Historically, Drosophila melanogaster has been the most extensively studied species in this family, but recent determination of the genomic sequences of an additional 11 Drosophila species opens up new avenues of research for other Drosophila species. This extensive sequence resource, encompassing species with well-defined phylogenetic relationships, provides a model system for comparative genomic analyses. FlyBase has developed tools to facilitate access to and navigation through this invaluable new data collection.

Animals↗

PubMed2006

Dynamics of DNA ejection from bacteriophage.

The ejection of DNA from a bacterial virus (i.e., phage) into its host cell is a biologically important example of the translocation of a macromolecular chain along its length through a membrane. The simplest mechanism for this motion is diffusion, but in the case of phage ejection a significant driving force derives from the high degree of stress to which the DNA is subjected in the viral capsid. The translocation is further sped up by the ratcheting and entropic forces associated with proteins that bind to the viral DNA in the host cell cytoplasm. We formulate a generalized diffusion equation that includes these various pushing and pulling effects and make estimates of the corresponding speedups in the overall translocation process. Stress in the capsid is the dominant factor throughout early ejection, with the pull due to binding particles taking over at later stages. Confinement effects are also investigated, in the case where the phage injects its DNA into a volume comparable to the capsid size. Our results suggest a series of in vitro experiments involving the ejection of DNA into vesicles filled with varying amounts of binding proteins from phage whose state of stress is controlled by ambient salt conditions or by tuning genome length.

Bacteriophage lambda↗

PubMed2006

The effect of genome length on ejection forces in bacteriophage lambda.

A variety of viruses tightly pack their genetic material into protein capsids that are barely large enough to enclose the genome. In particular, in bacteriophages, forces as high as 60 pN are encountered during packaging and ejection, produced by DNA bending elasticity and self-interactions. The high forces are believed to be important for the ejection process, though the extent of their involvement is not yet clear. As a result, there is a need for quantitative models and experiments that reveal the nature of the forces relevant to DNA ejection. Here, we report measurements of the ejection forces for two different mutants of bacteriophage lambda, lambdab221cI26 and lambdacI60, which differ in genome length by approximately 30%. As expected for a force-driven ejection mechanism, the osmotic pressure at which DNA release is completely inhibited varies with the genome length: we find inhibition pressures of 15 atm and 25 atm, for the short and long genomes, respectively, values that are in agreement with our theoretical calculations.

Bacteriophage lambda↗

PubMed2005

Elasticity theory and shape transitions of viral shells.

Recently, continuum elasticity theory has been applied to explain the shape transition of icosahedral viral capsids--single-protein-thick crystalline shells--from spherical to "buckled" or faceted as their radius increases through a critical value determined by the competition between stretching and bending energies of a closed two-dimensional (2D) elastic network. In the present work we generalize this approach to capsids with nonicosahedral symmetries, e.g., spherocylindrical and conical shells. One key additional physical ingredient is the role played by nonzero spontaneous curvature. Another is associated with the special way in which the energy of the 12 topologically required fivefold sites depends on the "background" local curvature of the shell in which they are embedded. Systematic evaluation of these contributions leads to a shape "phase" diagram in which transitions are observed from icosahedral to spherocylindrical capsids as a function of the ratio of stretching to bending energies and of the spontaneous curvature of the 2D protein network. We find that the transition from icosahedral to spherocylindrical symmetry is continuous or weakly first order near the onset of buckling, leading to extensive shape degeneracy. These results are discussed in the context of experimentally observed variations in the shapes of a variety of viral capsids.

Capsid↗

PubMed2005

A genetic screen for maternal-effect suppressors of decapentaplegic identifies the eukaryotic translation initiation factor 4A in Drosophila.

The Dpp signaling pathway is essential for many developmental processes in Drosophila and its activity is tightly regulated. To identify additional regulators of Dpp signaling, we conducted a genetic screen for maternal-effect suppressors of dpp haplo-insufficiency. We screened approximately 7000 EMS-mutagenized genomes and isolated and mapped seven independent dominant suppressors of dpp, Su(dpp), which were recovered as second-site mutations that resulted in viable flies in trans-heterozygous with dpp(H46), a dpp null allele. Most of the Su(dpp) mutants exhibited increased cell numbers of the amnioserosa, a cell type specified by the Dpp pathway, suggesting that these mutations may augment Dpp signaling activity. Here we report the unexpected identification of one of the Su(dpp) mutations as an allele of the eukaryotic translation initiation factor 4A (eIF4A). We show that Su(dpp)(YE9) maps to eIF4A and that this allele is associated with a substitution, arginine 321 to histidine, at a well-conserved amino acid and behaves genetically as a dominant-negative mutation. This result provides an intriguing link between a component of the translation machinery and Dpp signaling.

Amino Acid Substitution↗

PubMed2005

Structural polymorphism of the cytoskeleton: a model of linker-assisted filament aggregation.

The phase behavior of charged rods in the presence of interrod linkers is studied theoretically as a model for the equilibrium behavior underlying the organization of actin filaments by linker proteins in the cytoskeleton. The presence of linkers in the solution modifies the effective interrod interaction and can lead to interfilament attraction. Depending on the composition and physical properties of the system, such as linker-binding energies, filaments will orient either perpendicular or parallel to each other, leading to network-like or bundled structures. We show that such a system can have one of three generic phase diagrams, one dominated by bundles, another by networks, and the third containing both bundle and network-like phases. The first two diagrams can be found over a wide range of interaction energies, whereas the third diagram occurs only for a narrow range. These results provide theoretical understanding of the classification of linker proteins as bundling proteins or crosslinking proteins. In addition, they suggest possible mechanisms by which the cell may control cytoskeletal morphology.

Actins↗

PubMed2005

Research resources for Drosophila: the expanding universe.

Drosophila melanogaster has been the subject of research into central questions about biological mechanisms for almost a century. The experimental tools and resources that are available or under development for D. melanogaster and its related species, particularly those for genomic analysis, are truly outstanding. Here we review three types of resource that have been developed for D. melanogaster research: databases and other sources of information, biological materials and experimental services. These resources are there to be exploited and we hope that this guide will encourage new uses for D. melanogaster information, materials and services, both by those new to flies and by experienced D. melanogaster researchers.

Animals↗

PubMed2004

Measurements of DNA lengths remaining in a viral capsid after osmotically suppressed partial ejection.

The effect of external osmotic pressure on the extent of DNA ejection from bacteriophage-lambda was recently investigated (Evilevitch et al., 2003). The total length of DNA ejected was measured via the 260-nm absorption by free nucleotides, after opening of the capsids in the presence of varying amounts of polyethylene glycol 8000 and DNase I. As a function of osmolyte concentration, this absorption was shown to decrease progressively, ultimately vanishing completely for a sufficiently high external osmotic pressure. In this work we report the results of both sedimentation and gel analysis of the length of DNA remaining inside the capsids, as a function of osmolyte concentration. It is confirmed in this way that the progressive inhibition of DNA ejection corresponds to partial ejection from all of the capsids.

Absorption↗

PubMed2004

Origin of icosahedral symmetry in viruses.

With few exceptions, the shells (capsids) of sphere-like viruses have the symmetry of an icosahedron and are composed of coat proteins (subunits) assembled in special motifs, the T-number structures. Although the synthesis of artificial protein cages is a rapidly developing area of materials science, the design criteria for self-assembled shells that can reproduce the remarkable properties of viral capsids are only beginning to be understood. We present here a minimal model for equilibrium capsid structure, introducing an explicit interaction between protein multimers (capsomers). Using Monte Carlo simulation we show that the model reproduces the main structures of viruses in vivo (T-number icosahedra) and important nonicosahedral structures (with octahedral and cubic symmetry) observed in vitro. Our model can also predict capsid strength and shed light on genome release mechanisms.

Biophysical Phenomena↗

PubMed2004

Effect of mono- and multivalent salts on angle-dependent attractions between charged rods.

Using molecular dynamics simulations we examine the effective interactions between two like-charged rods as a function of angle and separation. In particular, we determine how the competing electrostatic repulsions and multivalent-ion-induced attractions depend upon concentrations of simple and multivalent salts. We find that with increasing multivalent salt, the stable configuration of two rods evolves from isolated rods to aggregated perpendicular rods to aggregated parallel rods; at sufficiently high concentration, additional multivalent salt reduces the attraction. Monovalent salt enhances the attraction near the onset of aggregation and reduces it at a higher concentration of multivalent salt.

Journal Article↗

PubMed2004

A statistical-thermodynamic model of viral budding.

We present a simple statistical thermodynamic model for budding of viral nucleocapsids at the cell membrane. The membrane is modeled as a flexible lipid bilayer embedding linker (spike) proteins, which serve to anchor and thus wrap the membrane around the viral capsids. The free energy of a single bud is expressed as a sum of the bending energy of its membrane coat, the spike-mediated capsid-membrane adhesion energy, and the line energy associated with the bud's rim, all depending on the extent of wrapping (i.e., bud size), and density of spikes in the curved membrane. This self-energy is incorporated into a simple free energy functional for the many-bud system, allowing for different spike densities, and hence entropy, in the curved (budding) and planar membrane regions, as well as for the configurational entropy of the polydisperse bud population. The equilibrium spike densities in the coexisting, curved and planar, membrane regions are calculated as a function of the membrane bending energy and the spike-mediated adhesion energy, for different spike and nucleocapsid concentrations in the membrane plane, as well as for several values of the bud's rim energy. We show that complete budding (full wrapping of nucleocapsids) can only take place if the adhesion energy exceeds a certain, critical, bending free energy. Whenever budding takes place, the spike density in the mature virions is saturated, i.e., all spike adhesion sites are occupied. The rim energy plays an important role in determining the size distribution of buds. The fraction of fully wrapped buds increases as this energy increases, resulting eventually in an all-or-nothing mechanism, whereby nucleocapsids at the plasma membrane are either fully enveloped or completely naked (just touching the membrane). We also find that at low concentrations all capsids arriving at the membrane get tightly and fully enveloped. Beyond a certain concentration, corresponding approximately to a stoichiometric spike/capsid ratio, newly arriving capsids cannot be fully wrapped; i.e., the budding yield decreases.

Algorithms↗

PubMed2003

Osmotic pressure inhibition of DNA ejection from phage.

Bacterial viral capsids in aqueous solution can be opened in vitro by addition of their specific receptor proteins, with consequent full ejection of their genomes. We demonstrate that it is possible to control the extent of this ejection by varying the external osmotic pressure. In the particular case of bacteriophage lambda, the ejection is 50% inhibited by osmotic pressures (of polyethylene glycol) comparable to those operative in the cytoplasm of host bacteria; it is completely suppressed by a pressure of 20 atmospheres. Furthermore, our experiments monitor directly a dramatic decrease of the stress inside the unopened phage capsid upon addition of polyvalent cations to the host solution, in agreement with many recent theories of DNA interactions.

Bacteriophages↗

PubMed2003

What drives the translocation of stiff chains?

We study the dynamics of the passage of a stiff chain through a pore into a cell containing particles that bind reversibly to it. Using Brownian molecular dynamics simulations we investigate the mean first-passage time as a function of the length of the chain inside for different concentrations of binding particles. As a consequence of the interactions with these particles, the chain experiences a net force along its length whose calculated value from the simulations accounts for the velocity at which it enters the cell. This force can in turn be obtained from the solution of a generalized diffusion equation incorporating an effective Langmuir adsorption free energy for the chain plus binding particles. These results suggest a role of binding particles in the translocation process that is in general quite different from that of a Brownian ratchet. Furthermore, nonequilibrium effects contribute significantly to the dynamics; e.g., the chain often enters the cell faster than particle binding can be saturated, resulting in a force several times smaller than the equilibrium value.

Binding Sites↗

PubMed2003

Viral self-assembly as a thermodynamic process.

The protein shells, or capsids, of nearly all spherelike viruses adopt icosahedral symmetry. In the present Letter, we propose a statistical thermodynamic model for viral self-assembly. We find that icosahedral symmetry is not expected for viral capsids constructed from structurally identical protein subunits and that this symmetry requires (at least) two internal "switching" configurations of the protein. Our results indicate that icosahedral symmetry is not a generic consequence of free energy minimization but requires optimization of internal structural parameters of the capsid proteins.

Capsid↗

PubMed2003

Osmotic shock and the strength of viral capsids.

Osmotic shock is a familiar means for rupturing viral capsids and exposing their genomes intact. The necessary conditions for providing this shock involve incubation in high-concentration salt solutions, and lower permeability of the capsids to salt ions than to water molecules. We discuss here how values of the capsid strength can be inferred from calculations of the osmotic pressure differences associated with measured values of the critical concentration of incubation solution.

Capsid↗

PubMed2003

Forces and pressures in DNA packaging and release from viral capsids.

In a previous communication (Kindt et al., 2001) we reported preliminary results of Brownian dynamics simulation and analytical theory which address the packaging and ejection forces involving DNA in bacteriophage capsids. In the present work we provide a systematic formulation of the underlying theory, featuring the energetic and structural aspects of the strongly confined DNA. The free energy of the DNA chain is expressed as a sum of contributions from its encapsidated and released portions, each expressed as a sum of bending and interstrand energies but subjected to different boundary conditions. The equilibrium structure and energy of the capsid-confined and free chain portions are determined, for each ejected length, by variational minimization of the free energy with respect to their shape profiles and interaxial spacings. Numerical results are derived for a model system mimicking the lambda-phage. We find that the fully encapsidated genome is highly compressed and strongly bent, forming a spool-like condensate, storing enormous elastic energy. The elastic stress is rapidly released during the first stage of DNA injection, indicating the large force (tens of pico Newtons) needed to complete the (inverse) loading process. The second injection stage sets in when approximately 1/3 of the genome has been released, and the interaxial distance has nearly reached its equilibrium value (corresponding to that of a relaxed torus in solution); concomitantly the encapsidated genome begins a gradual morphological transformation from a spool to a torus. We also calculate the loading force, the average pressure on the capsid's walls, and the anisotropic pressure profile within the capsid. The results are interpreted in terms of the (competing) bending and interaction components of the packing energy, and are shown to be in good agreement with available experimental data.

Bacteriophage lambda↗

PubMed2002

Annotation of the Drosophila melanogaster euchromatic genome: a systematic review.

BACKGROUND: The recent completion of the Drosophila melanogaster genomic sequence to high quality and the availability of a greatly expanded set of Drosophila cDNA sequences, aligning to 78% of the predicted euchromatic genes, afforded FlyBase the opportunity to significantly improve genomic annotations. We made the annotation process more rigorous by inspecting each gene visually, utilizing a comprehensive set of curation rules, requiring traceable evidence for each gene model, and comparing each predicted peptide to SWISS-PROT and TrEMBL sequences. RESULTS: Although the number of predicted protein-coding genes in Drosophila remains essentially unchanged, the revised annotation significantly improves gene models, resulting in structural changes to 85% of the transcripts and 45% of the predicted proteins. We annotated transposable elements and non-protein-coding RNAs as new features, and extended the annotation of untranslated (UTR) sequences and alternative transcripts to include more than 70% and 20% of genes, respectively. Finally, cDNA sequence provided evidence for dicistronic transcripts, neighboring genes with overlapping UTRs on the same DNA sequence strand, alternatively spliced genes that encode distinct, non-overlapping peptides, and numerous nested genes. CONCLUSIONS: Identification of so many unusual gene models not only suggests that some mechanisms for gene regulation are more prevalent than previously believed, but also underscores the complex challenges of eukaryotic gene prediction. At present, experimental data and human curation remain essential to generate high-quality genome annotations.

Animals↗