Jcb_200809046 323..340

Dendritic spines, small actin-rich protrusions from dendritic shafts, are the primary locus of excitatory synapses on neurons. Changes in dendritic spine morphology play a key role in memory formation and learning (Kasai et al., 2003). The loss or malformation of spines is also linked to many neurological diseases, which indicates the importance of proper regulation of spine morphology (Calabrese et al., 2006). Spines come in a wide range of sizes and shapes, even within the same brain region and the same dendrite. Individual spines also change shape continuously. Developmental shape changes follow a progressive replacement of the thin, elongated, and highly motile filopodia-like structures by more stable dendritic spines, which reach morphological maturity with a distinct neck and head (Oray et al., 2006). The actin cytoskeleton is central to numerous cellular processes involving membrane dynamics such as motility, morphogenesis, and endocytosis. During these processes, the barbed ends of polymerizing actin filaments push the membrane and promote the formation of plasma membrane protrusions or invaginations (Pollard and Borisy, 2003; Kaksonen et al., 2006; Carlier and Pantaloni, 2007). The actin cytoskeleton also plays a pivotal role in dendritic spine morphogenesis and dynamics (Ethell and Pasquale, 2005; Sekino et al., 2007; Zhang and Macara, 2008). Numerous actin-binding proteins under the control of different signaling pathways strictly regulate the dynamics of actin filaments, but information about the role of these proteins in dendritic spine morphogenesis is limited. The Arp2/3 complex is probably the most thoroughly characterized actin regulator in spine morphogenesis. Arp2/3 promotes nucleation of a branched actin filament network in other cell types (Pollard and Borisy, 2003; Pollard, 2007), and its expression is important for spine and synapse density (Wegner et al., 2008). Furthermore, reduced expression of its regulators affects spine morphology (Grove et al., 2004; Kim et al., 2006; Proepper et al., 2007; Soderling et al., 2007). Although these studies suggest that Arp2/3 complex induces formation of the branched actin network in the bulbous spine head, experimental evidence is still lacking. Dendritic spines are small protrusions along dendrites where the postsynaptic components of most excitatory synapses reside in the mature brain. Morphological changes in these actin-rich structures are associated with learning and memory formation. Despite the pivotal role of the actin cytoskeleton in spine morphogenesis, little is known about the mechanisms regulating actin filament polymerization and depolymerization in dendritic spines. We show that the filopodialike precursors of dendritic spines elongate through actin polymerization at both the filopodia tip and root. The small GTPase Rif and its effector mDia2 formin play a central role in regulating actin dynamics during filopodia elongation. Actin filament nucleation through the Arp2/3 complex subsequently promotes spine head expansion, and ADF/cofilin-induced actin filament disassembly is required to maintain proper spine length and morphology. Finally, we show that perturbation of these key steps in actin dynamics results in altered synaptic transmission. Defining mechanisms of actin polymerization and depolymerization during dendritic spine morphogenesis