Mesoscale model of postsynaptic density proteins.

(A) Schematics and molecular representation of the four PSD components in this simulation study. AMPAR and NMDAR diffuse on membrane, N-terminal part of PSD-95 is embedded on membrane due to the palmitoylation, and CaMKIIα is present beneath the membrane. (B) Three pairs of molecules with specific interactions between those domains. PDZ domains in PSD-95 can interact with PDZ binding motif (PBM) in TARP (top left) and with PBM in C-terminus of GluN2B subunit (top right). Only active CaMKII can interact with GluN2Bc by its kinase domains (bottom). Interaction strengths between all pair of molecules are set to reproduce the Kd values obtained from previous studies. (C) The 3D system simulation box. Core spherical particles of AMPAR and NMDAR are substituted by fluorescent proteins DsRed2 and eqFP670, respectively. (D) The 2D system setup and simulation box. Particles representing AMPAR, NMDAR, and N-terminal of PSD-95 diffuse on the planar membrane (orange), whereas other particles diffuse in the cytosol beneath the membrane (sky-blue).

Trajectories and morphologies in simulations of four-component PSD systems.

(A,B) Representative trajectories in the 3D (A) and 2D (B) systems. Bar chart to the right of the trajectory shows the average molecular compositions of AMPARs (A, red), NMDARs (N, orange), PSD-95s (P, blue) and CaMKIIs (C, green) in the largest cluster in the final 1000 snapshots. Dashed horizontal lines represent the whole number of proteins in the system. (C, E) Overview (left) and cross-sectional view (right) final snapshots of simulations in the 3D system with all CaMKIIs in the activated state (C) and in the inactive state (E). (D, F) Density profiles in the slice of the droplet along z axis (left) and the molecular distribution as a function of the distance from the center of mass of the cluster (right) with all CaMKIIs in the active state (D) and in the inactive state (F). (G, I) Top view (left) and side view (right) snapshots of simulations in the 2D system with CaMKII in the active (G) and inactive (I) states. (H, J) Density profiles in the slice of the cluster on membrane along x axis (left) and molecular distribution on membrane along the distance from the center of mass of the cluster (right) with all CaMKIIs in the active state (H) and in the inactive state (J). Shaded areas indicate standard errors for three independent simulations. The colors of the molecules, trajectories, bar graphs and distributions are corresponding.

Molecular interaction and CaMKII architecture in the four-component PSD assembly.

(A) Number of specific interaction (top) and non-specific interaction (bottom) with active CaMKII in 3D (blue), inactive CaMKII in 3D (sky-blue) active CaMKII in 2D (orange) and inactive CaMKII in 2D (pale orange). Each index indicates the interaction type where A, P, N, and C represent AMPAR, PSD-95, NMDAR, and CaMKII, respectively. (B) Distribution of the number of NMDARs bound to a CaMKII molecules in 3D system (top) and 2D system (bottom). (C) Protein distribution normal to the membrane plane (the z-coordinate) in the 2D system with the active (left) and inactive CaMKIIs (right). The orange and light blue background represent the membrane and cytoplasmic regions, respectively. (D) Two-dimensional kernel density estimation plot of the position and orientation of CaMKII molecules. The horizontal axis represents the distance from the membrane, while the vertical axis shows the angle between the membrane plane and each two hexagonal planes of CaMKII kinases. The height of each histogram and the color of KDE plot are shown in logarithmic scale. (E) Representative snapshots of the 2D systems with active CaMKII. The hexagonal plane of CaMKII is parallel to the membrane plane (left) or vertical to the membrane plane (middle). A snapshot of a whole cluster looking from the cytoplasm (right). Orange plane corresponds to the membrane center (z = 0).

Multiphase morphology for CaMKII with reduced volume in the 3D system.

(A) Schematics of CaMKIIs used in each simulation. Original sized CaMKII (WT) (left), CaMKII (r=2/3) with 2/3 of its radius (middle), and CaMKII (r=1/2) with 1/2 of its radius (right). (B) Snapshots of cross-sectional view with original CaMKII (WT) (left), CaMKII (r=2/3) (middle), and CaMKII (r=1/2) (right). (C) Density profile of AMPAR (red) and NMDAR (orange) in the slice of the cluster along z axis for CaMKII (WT) (left), CaMKII (r=2/3) (middle), and CaMKII (r=1/2) (right). (D) Molecular distribution of AMPAR (red) and NMDAR (orange) from the center of mass of the cluster (right) of the simulation with CaMKII (WT) (top), that with 2/3 in its radius (middle), and with 1/2 in its radius (bottom). As the molecular radius decreases, peak of AMPARs shifted outside while that of NMDARs moved close to the cluster center. (E) Molecular composition of each phases in the simulation with CaMKII (WT) (top), CaMKII (r=2/3) (middle), and CaMKII (r=1/2) (bottom). In each case, the three bars indicate the numbers of molecules in the AMPAR-containing phase (top bar), the NMDAR-containing phase (middle bar) and the dilute phase (bottom bar).

Multiphase morphology for CaMKII with reduced valency in the 2D system.

(A) Schematics of CaMKII used in each simulation. Original 12 valence CaMKII (WT) (left), 6 valence CaMKII (V=6) (middle), and 3 valence CaMKII (V=3) (right). (B) Snapshots of cross-sectional view with CaMKII (WT) (left), that with CaMKII (V=6) (middle), and with CaMKII (V=3) (right). (C) Density profile of AMPAR (red) and NMDAR (orange) in the slice of the cluster along Z axis of the simulation with CaMKII (WT) (left), CaMKII (V=6) (middle), and CaMKII (V=3) (right). As the valency decreases, the architecture of the core-shell cluster shifts between AMPARs and NMDARs. (D) Molecular distribution of AMPAR (red) and NMDAR (orange) from the center of mass of the cluster (right) of the simulation with CaMKII (WT) (top), that with CaMKII (V=6) (top middle), that with CaMKII (V=3) (bottom middle), and that with inactive CaMKII (bottom). As the molecular valence decreases, peak of AMPARs shifts inside while that of NMDARs moves far from the cluster center. (E) Molecular composition of each cluster in the simulation with CaMKII (WT) (top), that with CaMKII (V=6) (top middle), that with CaMKII (V=3) (bottom middle), and that with inactive CaMKII (bottom). The three bars indicate the number of molecules in each of the phases containing AMPARs (top bar), NMDARs (middle bar) and others (bottom bar), respectively. Composition of phase with NMDAR does not change drastically between different CaMKII sizes.

Interfacial tensions explain the multiphase morphology in 3D system.

(A) Representative snapshots of 3D slab simulation used in the measurement of interfacial tension between A (AMPAR-containing, red) and N (NMDAR-containing, green) phases, with CaMKII (WT) (top), CaMKII (r=2/3) (middle), and CaMKII (r=1/2) (bottom). (B) Interfacial tensions between the A and dilute phases (γA, red line), between the N and dilute phases (γN, orange line) and between the A and N phases (γAN, cyan line) with error bars at each condition. (C) Phase diagram on the γA⁄γAN and γN⁄γAN plane. Dashed lines represent theoretical borders of multiphase morphology.