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Kelley, C. F.

Publications and source records attributed to Kelley, C. F..

2 recordsLinked to original sources

Reconstitution of contractile actomyosin rings in vesicles

One of the grand challenges of bottom-up synthetic biology is the development of minimal machineries for cell division. The mechanical transformation of large-scale compartments, such as Giant Unilamellar Vesicles (GUVs), requires the geometry-specific coordination of active elements, several orders of magnitude larger than the molecular scale. Of all cytoskeletal structures, large-scale actomyosin rings appear to be the most promising cellular elements to accomplish this task. Here, we have adopted advanced encapsulation methods to study bundled actin filaments in GUVs and compare our results with theoretical modeling. By changing few key parameters, actin polymerization can be differentiated to resemble various types of networks in living cells. Importantly, we find membrane binding to be crucial for the robust condensation into a single actin ring in spherical vesicles, as predicted by theoretical considerations. Upon force generation by ATP-driven myosin motors, these ring-like actin structures contract and locally constrict the vesicle, forming furrow-like deformations. On the other hand, cortex-like actin networks are shown to induce and stabilize deformations from spherical shapes.

synthetic biology

Autoregulation clamps the synaptic membrane-remodeling machinery and promotes productive actin-dependent endocytosis

Synaptic membrane-remodeling events such as endocytosis require force-generating actin assembly. The endocytic machinery that regulates these actin and membrane dynamics localizes at high concentrations to large areas of the presynaptic membrane, but actin assembly and productive endocytosis are far more restricted in space and time. Here we describe a mechanism whereby autoinhibition clamps the presynaptic endocytic machinery to limit actin assembly to discrete functional events. We found that collective interactions between the Drosophila endocytic proteins Nwk/FCHSD2, Dap160/Intersectin, and WASp relieve Nwk autoinhibition and promote robust membrane-coupled actin assembly in vitro. Using automated particle tracking to quantify synaptic actin dynamics in vivo, we discovered that Nwk-Dap160 interactions constrain spurious assembly of WASp-dependent actin structures. These interactions also promote synaptic endocytosis, suggesting that autoinhibition both clamps and primes the synaptic endocytic machinery, thereby constraining actin assembly to drive productive membrane remodeling in response to physiological cues.

cell biology