SynGAP is a Ras/Rap GTPase-activating protein (GAP) that is a major constituent of postsynaptic densities (PSDs) from mammalian forebrain. Its α1 isoform binds to all three PDZ (PSD-95, Discs-large, ZO-1) domains of PSD-95, the principal PSD scaffold, and can occupy as many as 15% of these PDZ domains. We present evidence that synGAP-α1 regulates the composition of the PSD by restricting binding to the PDZ domains of PSD-95. We show that phosphorylation by Ca2+/calmodulin-dependent protein kinase II (CaMKII) and Polo-like kinase-2 (PLK2) decreases its affinity for the PDZ domains by several fold, which would free PDZ domains for occupancy by other proteins. Finally, we show that three critical postsynaptic signaling proteins that bind to the PDZ domains of PSD-95 are present in higher concentration in PSDs isolated from mice with a heterozygous deletion of synGAP.DOI: http://dx.doi.org/10.7554/eLife.16813.001

Calmodulin-dependent kinase II (CaMKII) has long been known to play an important role in learning and memory as well as long term potentiation (LTP). More recently it has been suggested that it might be involved in the time averaging of synaptic signals, which can then lead to the high precision of information stored at a single synapse. However, the role of the scaffolding molecule, neurogranin (Ng), in governing the dynamics of CaMKII is not yet fully understood. In this work, we adopt a rule-based modeling approach through the Monte Carlo method to study the effect of Ca 2+ signals on the dynamics of CaMKII phosphorylation in the postsynaptic density (PSD). Calcium surges are observed in synaptic spines during an EPSP and back-propagating action potential due to the opening of NMDA receptors and voltage dependent calcium channels. Using agent-based models, we computationally investigate the dynamics of phosphorylation of CaMKII monomers and dodecameric holoenzymes. The scaffolding molecule, Ng, when present in significant concentration, limits the availability of free calmodulin (CaM), the protein which activates CaMKII in the presence of calcium. We show that Ng plays an important modulatory role in CaMKII phosphorylation following a surge of high calcium concentration. We find a non-intuitive dependence of this effect on CaM concentration that results from the different affinities of CaM for CaMKII depending on the number of calcium ions bound to the former. It has been shown previously that in the absence of phosphatase, CaMKII monomers integrate over Ca 2+ signals of certain frequencies through autophosphorylation (Pepke et al, Plos Comp. Bio., 2010). We also study the effect of multiple calcium spikes on CaMKII holoenzyme autophosphorylation, and show that in the presence of phosphatase, CaMKII behaves as a leaky integrator of calcium signals, a result that has been recently observed in vivo. Our models predict that the parameters of this leaky integrator are finely tuned through the interactions of Ng, CaM, CaMKII, and PP1, providing a mechanism to precisely control the sensitivity of synapses to calcium signals.

Voltage-gated Cav1.2 and Cav1.3 (L-type) Ca2+channels regulate neuronal excitability, synaptic plasticity, and learning and memory. Densin-180 (densin) is an excitatory synaptic protein that promotes Ca2+-dependent facilitation of voltage-gated Cav1.3 Ca2+channels in transfected cells. Mice lacking densin (densin KO) exhibit defects in synaptic plasticity, spatial memory, and increased anxiety-related behaviors—phenotypes that more closely match those in mice lacking Cav1.2 than Cav1.3. Therefore, we investigated the functional impact of densin on Cav1.2. We report that densin is an essential regulator of Cav1.2 in neurons, but has distinct modulatory effects compared with its regulation of Cav1.3. Densin binds to the N-terminal domain of Cav1.2, but not that of Cav1.3, and increases Cav1.2 currents in transfected cells and in neurons. In transfected cells, densin accelerates the forward trafficking of Cav1.2 channels without affecting their endocytosis. Consistent with a role for densin in increasing the number of postsynaptic Cav1.2 channels, overexpression of densin increases the clustering of Cav1.2 in dendrites of hippocampal neurons in culture. Compared with wild-type mice, the cell surface levels of Cav1.2 in the brain, as well as Cav1.2 current density and signaling to the nucleus, are reduced in neurons from densin KO mice. We conclude that densin is an essential regulator of neuronal Cav1 channels and ensures efficient Cav1.2 Ca2+signaling at excitatory synapses.SIGNIFICANCE STATEMENTThe number and localization of voltage-gated CavCa2+channels are crucial determinants of neuronal excitability and synaptic transmission. We report that the protein densin-180 is highly enriched at excitatory synapses in the brain and enhances the cell surface trafficking and postsynaptic localization of Cav1.2 L-type Ca2+channels in neurons. This interaction promotes coupling of Cav1.2 channels to activity-dependent gene transcription. Our results reveal a mechanism that may contribute to the roles of Cav1.2 in regulating cognition and mood.

SynGAP is a postsynaptic density (PSD) protein that binds to PDZ domains of the scaffold protein PSD-95. We previously reported that heterozygous deletion of Syngap1 in mice is correlated with increased steady-state levels of other key PSD proteins that bind PSD-95, although the level of PSD-95 remains constant (Walkup et al., 2016). For example, the ratio to PSD-95 of Transmembrane AMPA-Receptor-associated Proteins (TARPs), which mediate binding of AMPA-type glutamate receptors to PSD-95, was increased in young Syngap1+/-mice. Here we show that only females and not males show a highly significant correlation between an increase in TARP and a decrease in synGAP in the PSDs of Syngap1+/-rodents. The data reveal a sex difference in the adaptation of the PSD scaffold to synGAP haploinsufficiency.

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