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Blanchoin, Laurent (Ed.)IQGAP is a conserved family of actin-binding proteins with essential roles in cell motility, cytokinesis, and cell adhesion, yet there remains a limited understanding of how IQGAP proteins directly influence actin filament dynamics. To close this gap, we used single-molecule and single-filament total internal reflection fluorescence microscopy to observe IQGAP regulating actin dynamics in real time. To our knowledge, this is the first study to do so. Our results demonstrate that full-length human IQGAP1 forms dimers that stably bind to actin filament sides and transiently cap barbed ends. These interactions organize filaments into thin bundles, suppress barbed end growth, and inhibit filament disassembly. Surprisingly, each activity depends on distinct combinations of IQGAP1 domains and/or dimerization, suggesting that different mechanisms underlie each functional effect on actin. These observations have important implications for how IQGAP functions as an actin regulator in vivo and how it may be regulated in different biological settings.more » « less
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null (Ed.)In many real-world scenarios, the time it takes for a mobile agent, e.g., a robot, to move from one location to another may vary due to exogenous events and be difficult to predict accurately. Planning in such scenarios is challenging, especially in the context of Multi-Agent Pathfinding (MAPF), where the goal is to find paths to multiple agents and temporal coordination is necessary to avoid collisions. In this work, we consider a MAPF problem with this form of time uncertainty, where we are only given upper and lower bounds on the time it takes each agent to move. The objective is to find a safe solution, which is a solution that can be executed by all agents and is guaranteed to avoid collisions. We propose two complete and optimal algorithms for finding safe solutions based on well-known MAPF algorithms, namely, A* with Operator Decomposition (A* + OD) and Conflict-Based Search (CBS). Experimentally, we observe that on several standard MAPF grids the CBS-based algorithm performs better. We also explore the option of online replanning in this context, i.e., modifying the agents' plans during execution, to reduce the overall execution cost. We consider two online settings: (a) when an agent can sense the current time and its current location, and (b) when the agents can also communicate seamlessly during execution. For each setting, we propose a replanning algorithm and analyze its behavior theoretically and empirically. Our experimental evaluation confirms that indeed online replanning in both settings can significantly reduce solution cost.more » « less
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Twinfilin bypasses assembly conditions and actin filament aging to drive barbed end depolymerization
Cellular actin networks grow by ATP-actin addition at filament barbed ends and have long been presumed to depolymerize at their pointed ends, primarily after filaments undergo “aging” (ATP hydrolysis and Pi release). The cytosol contains high levels of actin monomers, which favors assembly over disassembly, and barbed ends are enriched in ADP-Pi actin. For these reasons, the potential for a barbed end depolymerization mechanism in cells has received little attention. Here, using microfluidics-assisted TIRF microscopy, we show that mouse twinfilin, a member of the ADF-homology family, induces depolymerization of ADP-Pi barbed ends even under assembly-promoting conditions. Indeed, we observe in single reactions containing micromolar concentrations of actin monomers the simultaneous rapid elongation of formin-bound barbed ends and twinfilin-induced depolymerization of free barbed ends. The data show that twinfilin catalyzes dissociation of subunits from ADP-Pi barbed ends and thereby bypasses filament aging prerequisites to disassemble newly polymerized actin filaments.
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Abstract Cellular actin networks can be rapidly disassembled and remodeled in a few seconds, yet in vitro actin filaments depolymerize slowly over minutes. The cellular mechanisms enabling actin to depolymerize this fast have so far remained obscure. Using microfluidics-assisted TIRF, we show that Cyclase-associated protein (CAP) and Cofilin synergize to processively depolymerize actin filament pointed ends at a rate 330-fold faster than spontaneous depolymerization. Single molecule imaging further reveals that hexameric CAP molecules interact with the pointed ends of Cofilin-decorated filaments for several seconds at a time, removing approximately 100 actin subunits per binding event. These findings establish a paradigm, in which a filament end-binding protein and a side-binding protein work in concert to control actin dynamics, and help explain how rapid actin network depolymerization is achieved in cells.
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Mapping dislocation densities resulting from severe plastic deformation using large strain machiningThe multiplication of dislocations determines the trajectories of microstructure evolution during plastic deformation. It has been recognized that the dislocation storage and the deformation-driven subgrain formation are correlated—the principle of similitude, where the dislocation density (ρ i ) scales self-similarly with the subgrain size (δ): $\delta \sqrt {{\rho _{\rm{i}}}}$ ∼ constant. Here, the robustness of this concept in Cu is probed utilizing large strain machining across a swathe of severe shear deformation conditions—strains in the range 1–10 and strain-rates 10–10 3 /s. Deformation strain, strain-rate, and temperature characterizations are juxtaposed with electron microscopy, and dislocation densities are measured by quantification of broadening of X-ray diffraction peaks of crystallographic planes. We parameterize the variation of dislocation density as a function of strain and a rate parameter R , a function of strain-rate, temperature, and material constants. We confirm the preservation of similitude between dislocation density and the subgrain structure across orders-of-magnitude of thermomechanical conditions.more » « less
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This article presents an overview of the collaborative Transit Hub project between Vanderbilt University, the Nashville Metropolitan Transit Authority (MTA) and Siemens, Corporate Technology. This project commenced as part of the NIST Global Cities Team Challenge (GCTC). The goal of this project is to leverage technology effectively to improve public engagement with transit operations and increase the overall efficiency of the system. In the process we want to identify key technical challenges that will require new research to advance the state of the art.more » « less