More than a mere supply of monomers: G-Actin pools regulate actin dynamics in dendritic spines

2255 The Rockefeller University Press J. Cell Biol. Vol. 216 No. 8 2255–2257 https://doi.org/10.1083/jcb.201705216 Dendritic spines are small, micrometer-sized actin-rich protrusions formed along the dendrites of excitatory and inhibitory neurons, which serve as the postsynaptic component for most of the excitatory inputs within the central nervous system. Dendritic spines are highly motile protrusions, which rapidly change their morphology and molecular composition according to synaptic activity. It is widely accepted that spine size and shape are intimately linked to synaptic plasticity. Thus, the strengthening or weakening of synaptic connections during long-term potentiation (LTP) or long-term depression are accompanied by the enlargement or shrinkage of dendritic spine heads, respectively (Okamoto et al., 2004). The major cytoskeletal element of dendritic spines is actin, which serves both as a structural and dynamic framework and as the principal regulator of protein and vesicular trafficking. The organization of filamentous actin (F-actin) is determined upon the balance between (a) the addition of monomeric globular actin (G-actin) to the growing, barbed ends, (b) the generation of side branches, (c) the severing or (d) stabilization of the existing filaments, and (e) the depolymerization at the pointed ends (Bosch et al., 2014). So far, most studies have concentrated on the formation and maintenance of the F-actin network within dendritic spines and less emphasis was given on the local availability of G-actin, which provides the monomer supply needed for filament formation. The intracellular concentration of G-actin exceeds with several orders of magnitude the critical concentration needed for rapid filament formation (Koestler et al., 2009), leading to a general assumption that the pool of G-actin provides a constant supply of monomers in a diffusible and excessive manner. In contrast, several evolutionary conserved actin-monomer binding proteins regulate the availability and subcellular localization as well as the nucleotide status of actin monomers (thus, ADP– G-actin or ATP–G-actin; Paavilainen et al., 2004). Therefore, it is highly likely that these proteins provide additional means of spatial and temporal regulation of actin polymerization. In this issue, Lei et al. report that the local enrichment of the G-actin pool plays an important role in regulating basal and activity-dependent actin polymerization within the dendritic spine. Working in primary dissociated and organotypic hippocampal cultures, they show that G-actin is enriched in dendritic spines under basal conditions as well as after chemically induced LTP (cLTP). To quantify the distribution of actin monomers between dendritic spines and the dendrite shaft, specific probes to endogenous G-actin (vitamin D–binding protein and an anti–G-actin antibody; Lee et al., 2013) and overexpression of EGFP-tagged, nonpolymerizable γ-actin mutants (R62D and G13R) were applied. In intact neurons, spine head-to-shaft ratios of fluorescent signals were elevated, suggesting a local enrichment of G-actin in spine heads. Signal intensity was reduced upon short-term permeabilization of living cells with the mild detergent saponin, confirming the specificity of detecting only nonpolymerized, monomer actin. The amount of EGFP-tagged actin monomers within the dendritic spine heads was increased rapidly upon TEA or glycine-induced cLTP, but the presence of nonpolymerizable γ-actin mutants inhibited spine head enlargement, known to occur during cLTP-induced structural plasticity (Bosch et al., 2014). Importantly, these data suggest that G-actin enrichment within the dendritic spines is regulated by synaptic activity but independently from actin polymerization. Careful FRAP assays in dendritic spines expressing EGFP-tagged wild-type or the R62D and G13R actin monomers revealed recovery curves that were all fitted with double exponential functions but had significantly different time constants of the recovery curves. Both actin mutants and wild-type actin recovery curves exhibited similar rapid components, indicating that the diffusion of the three EGFP-tagged actin proteins into the spines happens with similar kinetics. In contrast, the slow recovery rate of the mutant actin monomers was significantly delayed, suggesting the possibility that these mutants might get trapped within the spine heads after their rapid delivery. Superresolution imaging of actin flow and the distribution of proteins regulating actin polymerization within the spine already indicated distinct nanoscale domains for slow and nonpolarized actin nucleation in the close vicinity of the postsynaptic density or promoting finger-like protrusions at the perisynaptic sites via fast actin polymerization (Frost et al., 2010; Chazeau et al., 2014). This spatially and functionally distinct actin turnover Synaptic activity reshapes the morphology of dendritic spines via regulating F-actin arborization. In this issue, Lei et al. (2017. J. Cell Biol. https ://doi .org /10 .1083 / jcb .201612042) reports a novel, G-actin–dependent regulation of actin polymerization within spine heads. They show that actin monomer levels are elevated in spines upon activity, with G-actin immobilized by the local enrichment of phosphatidylinositol (3,4,5)-triphosphate (PIP3) within the spine plasma membrane. More than a mere supply of monomers: G-Actin pools regulate actin dynamics in dendritic spines