βCaMKII Regulates Actin Assembly and Structure

Abstract

Ca2؉ -Calmodulin-dependent protein kinase II (CaMKII) is an abundant synaptic protein that was recently shown to regulate the organization of actin filaments leading to structural modifications of synapses. CaMKII is a dodecameric complex with a special architecture that provides it with unique potential for organizing the actin cytoskeleton. We report using biochemical assays that the ␤ isoform of CaMKII binds to and bundles actin filaments, and the disposition of ␤CaMKII within the actin bundles was revealed by cryoelectron tomography. In addition, ␤CaMKII was found to inhibit actin polymerization, suggesting that it either serves as a capping protein or binds monomeric actin, reducing the amount of freely available monomers to nucleate polymer assembly. By means of fluorescent cross-correlation spectroscopy, we determined that ␤CaMKII does indeed bind to monomeric actin, reaching saturation at a stoichiometry of 12:1 actin monomers per ␤CaMKII holoenzyme with a binding constant of 2.4 ؋ 10 5 M ؊1 . In cells, ␤CaMKII has a dual functional role; it can sequester monomeric actin to reduce actin polymerization and can also bundle actin filaments. Together, these effects would impact both the dynamics of actin filament assembly and enhance the rigidity of the filaments once formed, significantly impacting the structure of synapses.Actin and more than 60 different classes of actin-binding proteins form an abundant and highly regulated cytoskeletal network, constituting more than 25% of protein in non-muscle cells and over 60% in muscle cells (1). Actin is estimated to be ϳ4 mg/ml in cells, with half found in the polymerized F-actin state and the remainder in the soluble G-actin state (2). Actin undergoes rounds of polymerization and depolymerization from a soluble (G-actin) pool to a filamentous (F-actin) pool, and it is the kinetics of these forward and backward rates that determines the stability of the actin cytoskeleton. The various actin-binding proteins are, in part, responsible for regulating the kinetics of actin polymerization and depolymerization. Actin is commonly known for its structural and dynamic function in cells, such as its role in muscle contraction and cell locomotion, and has been similarly implicated for its structural role at synapses in the nervous system (3, 4). For example, actin is proposed to serve an important role in coordinating the delivery of synaptic vesicles to the pre-synaptic terminal that appear to underlie certain forms of plasticity mediated by alterations in the probability of stimulus evoked vesicle release (5-7). Additionally, several recent studies have demonstrated the importance of actin dynamics in structural reorganization of the post-synaptic spine compartment of excitatory synapses (8 -10). Several pools of actin have been identified within spines with different turnover rates, and activation of glutamate receptors on the spines regulates the kinetics of actin turnover (9, 10). Spine shape changes have been proposed to underlie forms of synaptic plasticity such as l...

The Roles of CaMKII and F-Actin in the Structural Plasticity of Dendritic Spines: A Potential Molecular Identity of a Synaptic Tag?

“…This suggests that CaMKII hetero-oligomers might bind simultaneously to different actin filaments through multiple ␤-subunits and, thus, might confer on CaMKII the ability to bundle actin filaments together. Confirming this idea, F-actin has been found to forms thick bundled structures in vitro in the presence of the ␤-subunit of CaMKII but not in the presence of the ␣-subunit (FIGURE 2, C AND D) (93,96,110). Phosphorylation of CaMKII reduces this bundling activity, suggesting that this property can be regulated by synaptic activity (96).…”

Biochemical principles underlying the stable maintenance of LTP by the CaMKII/NMDAR complex

“…Holoenzymes having at least some beta subunits are bound to actin within the spine head at resting Ca 2+ levels (Okamoto et al, 2007; Sanabria et al, 2009; Shen et al, 1998). Upon binding of Ca 2+ /CaM, CaMKII is released from actin and can only then diffuse to the NMDAR.…”

CaMK-II is a PKD2 target that promotes pronephric kidney development and stabilizes cilia

“…Activated CaMK-II is enriched at the apical surface of ductal cells and is known to directly interact with and stabilize F-actin (Easley et al, 2006;Lin and Redmond, 2008;Okamoto et al, 2007;Sanabria et al, 2009). Apical constriction mediated by actinomyosin is necessary for the alterations in cell shape that lead to proper tissue development (Daggett et al, 2007), including duct formation (Sawyer et al, 2010) and could explain a role for CaMK-II in cloacal cells.…”