The bacterial pathogen Listeria monocytogenes propels itself in the cytoplasm of the infected cells by forming a filamentous comet tail assembled by the polymerization of the cytoskeletal protein actin. Although a great deal is known about the molecular processes that lead to actin-based movement, most macroscale aspects of motion, including the nature of the trajectories traced out by the motile bacteria, are not well understood. Here, we present 2D trajectories of Listeria moving between a glass-slide and coverslip in a Xenopus frog egg extract motility assay. We observe that the bacteria move in a number of fascinating geometrical trajectories, including winding S curves, translating figure eights, small-and largeamplitude sine curves, serpentine shapes, circles, and a variety of spirals. We then develop a dynamic model that provides a unified description of these seemingly unrelated trajectories. A key ingredient of the model is a torque (not included in any microscopic models of which we are aware) that arises from the rotation of the propulsive force about the body axis of the bacterium. We show that a large variety of trajectories with a rich mathematical structure are obtained by varying the rate at which the propulsive force moves about the long axis. The trajectories of bacteria executing both steady and saltatory motion are found to be in excellent agreement with the predictions of our dynamic model. When the constraints that lead to planar motion are removed, our model predicts motion along regular helical trajectories, observed in recent experiments. motility ͉ bacteria P olymerization of the cytoskeletal protein actin into a network of filaments is necessary for the motility of several infectious bacteria. These bacterial pathogens hijack the actin machinery of the host cell to form ''comet tails'' due to unidirectional actin assembly at one of their poles. The force generated by the actin network allows the bacteria to move within the infected cell and to other cells in it's neighborhood. The biochemistry of the tail formation process has been well studied in the case of the Grampositive bacterium Listeria monocytogenes (1-3). In particular, it has been shown that only one bacterial surface factor, ActA, is required for the movement of Listeria in a medium containing a few other actin-related proteins from the cytoplasm of the host cell. This observation has led to in vitro motility assays in which polystyrene beads (4) or disks (5) and phospholipid vesicles (6, 7) coated with ActA are propelled by the comet tails formed by actin polymerization. Many of the proteins responsible for the movement of Listeria have also been found in the front end of a crawling cell, also referred to as the lamellopodium (1). Listeria is therefore a model system that has provided important molecular level insights on actin-based motility.Although progress has been made at the molecular level, the connection between biochemical processes and force generation has not been fully elucidated. With the knowledge of polymeri...

Plant synthetic biology promises immense technological benefits, including the potential development of a sustainable bio-based economy through the predictive design of synthetic gene circuits. Such circuits are built from quantitatively characterized genetic parts; however, this characterization is a significant obstacle in work with plants because of the time required for stable transformation. We describe a method for rapid quantitative characterization of genetic plant parts using transient expression in protoplasts and dual luciferase outputs. We observed experimental variability in transient-expression assays and developed a mathematical model to describe, as well as statistical normalization methods to account for, this variability, which allowed us to extract quantitative parameters. We characterized >120 synthetic parts in Arabidopsis and validated our method by comparing transient expression with expression in stably transformed plants. We also tested >100 synthetic parts in sorghum (Sorghum bicolor) protoplasts, and the results showed that our method works in diverse plant groups. Our approach enables the construction of tunable gene circuits in complex eukaryotic organisms.

Using fluorescence microscopy, we directly visualize the condensed structures of individual semi-flexible actin filaments in a poor solvent. The condensation of filaments into either ring-like or racquet-like structures is driven by non-adsorbing polymers which induce attractive interactions between filaments via the well-known depletion mechanism. A quantitative analysis of the racquet structures yields a direct measurement of the adhesion strength between a pair of filaments. We also compare our experimental data with a theoretical model, demonstrating that in the limit of weak binding, thermal fluctuations can renormalize the effective strength of the attractive depletion interactions. Our experimental methods can be applied to other filamentous structures to directly measure their attractive intermolecular potentials.

Metastatic cancer cells for many cancers are known to have altered cytoskeletal properties, in particular to be more deformable and contractile. Consequently, shape characteristics of more metastatic cancer cells may be expected to have diverged from those of their parental cells. To examine this hypothesis we study shape characteristics of paired osteosarcoma cell lines, each consisting of a less metastatic parental line and a more metastatic line, derived from the former by in vivo selection. Two-dimensional images of four pairs of lines were processed. Statistical analysis of morphometric characteristics shows that shape characteristics of the metastatic cell line are partly overlapping and partly diverged from the parental line. Significantly, the shape changes fall into two categories, with three paired cell lines displaying a more mesenchymal-like morphology, while the fourth displaying a change towards a more rounded morphology. A neural network algorithm could distinguish between samples of the less metastatic cells from the more metastatic cells with near perfect accuracy. Thus, subtle changes in shape carry information about the genetic changes that lead to invasiveness and metastasis of osteosarcoma cancer cells.

T lymphocytes play a key role in adaptive immunity and are activated by interactions of their T cell receptors (TCR) with peptides (p) derived from antigenic proteins bound to MHC gene products. The repertoire of T lymphocytes available in peripheral organs is tuned in the thymus. Immature T lymphocytes (thymocytes) interact with diverse endogenous peptides bound to MHC in the thymus. TCR expressed on thymocytes must bind weakly to endogenous pMHC (positive selection) but must not bind too strongly to them (negative selection) to emerge from the thymus. Negatively selecting pMHC ligands bind TCR with a binding affinity that exceeds a sharply defined (digital) threshold. In contrast, there is no sharp threshold separating positively selecting ligands from those that bind too weakly to elicit a response. We describe results of computer simulations and experiments, which suggest that the contrast between the characters of the positive and negative selection thresholds originates in differences in the way in which Ras proteins are activated by ligands of varying potency. The molecular mechanism suggested by our studies generates hypotheses for how genetic aberrations may dampen the digital negative selection response, with concomitant escape of autoimmune T lymphocytes from the thymus.positive feedback ͉ Ras activation ͉ signal transduction ͉ T cell antigen receptor ͉ thymic development