Activity-dependent regulation of dendritic growth and maintenance by glycogen synthase kinase 3ß.

Activity-dependent dendritic development represents a crucial step in brain development, but its underlying mechanisms remain to be fully elucidated. Here we report that glycogen synthase kinase 3ß (GSK3ß) regulates dendritic development in an activity-dependent manner. We find that GSK3ß in somatodendritic compartments of hippocampal neurons becomes highly phosphorylated at serine-9 upon synaptogenesis. This phosphorylation-dependent GSK3ß inhibition is mediated by neurotrophin signalling and is required for dendritic growth and arbourization. Elevation of GSK3ß activity leads to marked shrinkage of dendrites, whereas its inhibition enhances dendritic growth. We further show that these effects are mediated by GSK3ß regulation of surface GABAA receptor levels via the scaffold protein gephyrin. GSK3ß activation leads to gephyrin phosphorylation to reduce surface GABAA receptor clusters, resulting in neuronal hyperexcitability that causes dendrite shrinkage. These findings thus identify GSK3ß as a key player in activity-dependent regulation of dendritic development by targeting the excitatory-inhibitory balance of the neuron.

Dynamic localization of G-actin during membrane protrusion in neuronal motility.

BACKGROUND: Actin-based cell motility is fundamental for development, function, and malignant events in eukaryotic organisms. During neural development, axonal growth cones depend on rapid assembly and disassembly of actin filaments (F-actin) for their guided extension to specific targets for wiring. Monomeric globular actin (G-actin) is the building block for F-actin but is not considered to play a direct role in spatiotemporal control of actin dynamics in cell motility. RESULTS: Here we report that a pool of G-actin dynamically localizes to the leading edge of growth cones and neuroblastoma cells to spatially elevate the G-/F-actin ratio that drives membrane protrusion and cell movement. Loss of G-actin localization leads to the cessation and retraction of membrane protrusions. Moreover, G-actin localization occurs asymmetrically in growth cones during attractive turning. Finally, we identify the actin monomer-binding proteins profilin and thymosin ß4 as key molecules that localize actin monomers to the leading edge of lamellipodia for their motility. CONCLUSIONS: Our results suggest that dynamic localization of G-actin provides a novel mechanism to regulate the spatiotemporal actin dynamics underlying membrane protrusion in cell locomotion and growth cone chemotaxis.

Instantaneous inactivation of cofilin reveals its function of F-actin disassembly in lamellipodia.

Cofilin is a key regulator of the actin cytoskeleton. It can sever actin filaments, accelerate filament disassembly, act as a nucleation factor, recruit or antagonize other actin regulators, and control the pool of polymerization-competent actin monomers. In cells these actions have complex functional outputs. The timing and localization of cofilin activity are carefully regulated, and thus global, long-term perturbations may not be sufficient to probe its precise function. To better understand cofilin's spatiotemporal action in cells, we implemented chromophore-assisted laser inactivation (CALI) to instantly and specifically inactivate it. In addition to globally inhibiting actin turnover, CALI of cofilin generated several profound effects on the lamellipodia, including an increase of F-actin, a rearward expansion of the actin network, and a reduction in retrograde flow speed. These results support the hypothesis that the principal role of cofilin in lamellipodia at steady state is to break down F-actin, control filament turnover, and regulate the rate of retrograde flow.