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, lambda b221cI26 and lambda cI60, which differ in genome length by ~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, respectively, values that are in agreement with our theoretical calculations

Measuring Flux Distributions for Diffusion in the Small-Numbers Limit

For the classical diffusion of independent particles, Fick's law gives a well-known relationship between the average flux and the average concentration gradient. What has not yet been explored experimentally, however, is the dynamical distribution of diffusion rates in the limit of small particle numbers. Here, we measure the distribution of diffusional fluxes using a microfluidics device filled with a colloidal suspension of a small number of microspheres. Our experiments show that (1) the flux distribution is accurately described by a Gaussian function; (2) Fick's law, that the average flux is proportional to the particle gradient, holds even for particle gradients down to a single particle difference; (3) the variance in the flux is proportional to the sum of the particle numbers; and (4) there are backward flows, where particles flow up a concentration gradient, rather than down it. In addition, in recent years, two key theorems about nonequilibrium systems have been introduced:  Evans' fluctuation theorem for the distribution of entropies and Jarzynski's work theorem. Here, we introduce a new fluctuation theorem, for the fluxes, and we find that it is confirmed quantitatively by our experiments

Broad-tailed force distributions and velocity ordering in a heterogeneous membrane model for collective cell migration

Correlated velocity patterns and associated large length-scale transmission of traction forces have been observed in collective live cell migration as a response to a "wound". We argue that a simple physical model of a force-driven heterogeneous elastic membrane sliding over a viscous substrate can qualitatively explain a few experimentally observed facts: (i) the growth of velocity ordering which spreads from the wound boundary to the interior, (ii) the exponential tails of the traction force distributions, and (iii) the swirling pattern of velocities in the interior of the tissue.Comment: 7 pages and 5 figure

Dynamics of DNA Ejection From Bacteriophage

The ejection of DNA from a bacterial virus (``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 speed-ups 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.Comment: 17 pages, 5 figure

Forces During Bacteriophage DNA Packaging and Ejection

The conjunction of insights from structural biology, solution biochemistry, genetics and single molecule biophysics has provided a renewed impetus for the construction of quantitative models of biological processes. One area that has been a beneficiary of these experimental techniques is the study of viruses. In this paper we describe how the insights obtained from such experiments can be utilized to construct physical models of processes in the viral life cycle. We focus on dsDNA bacteriophages and show that the bending elasticity of DNA and its electrostatics in solution can be combined to determine the forces experienced during packaging and ejection of the viral genome. Furthermore, we quantitatively analyze the effect of fluid viscosity and capsid expansion on the forces experienced during packaging. Finally, we present a model for DNA ejection from bacteriophages based on the hypothesis that the energy stored in the tightly packed genome within the capsid leads to its forceful ejection. The predictions of our model can be tested through experiments in vitro where DNA ejection is inhibited by the application of external osmotic pressure