Protein Kinase Cϵ (PKCϵ) Promotes Synaptogenesis through Membrane Accumulation of the Postsynaptic Density Protein PSD-95

Protein kinase C epsilon (PKC epsilon) promotes synaptic maturation and synaptogenesis via activation of synaptic growth factors such as BDNF, NGF, and IGF. However, many of the detailed mechanisms by which PKC epsilon induces synaptogenesis are not fully understood. Accumulation of PSD-95 to the postsynaptic density (PSD) is known to lead to synaptic maturation and strengthening of excitatory synapses. Here we investigated the relationship between PKC epsilon and PSD-95. We show that the PKC epsilon activators dicyclopropanated linoleic acid methyl ester and bryostatin 1 induce phosphorylation of PSD-95 at the serine 295 residue, increase the levels of PSD-95, and enhance its membrane localization. Elimination of the serine 295 residue in PSD-95 abolished PKC epsilon-induced membrane accumulation. Knockdown of either PKC epsilon or JNK1 prevented PKC epsilon activator-mediated membrane accumulation of PSD-95. PKC epsilon directly phosphorylated PSD-95 and JNK1 in vitro. Inhibiting PKC epsilon, JNK, or calcium/calmodulin-dependent kinase II activity prevented the effects of PKC epsilon activators on PSD-95 phosphorylation. Increase in membrane accumulation of PKC epsilon and phosphorylated PSD-95 (p-PSD-95(S295)) coincided with an increased number of synapses and increased amplitudes of excitatory post-synaptic potentials (EPSPs) in adult rat hippocampal slices. Knockdown of PKC epsilon also reduced the synthesis of PSD-95 and the presynaptic protein synaptophysin by 30 and 44%, respectively. Prolonged activation of PKC epsilon increased synapse number by 2-fold, increased presynaptic vesicle density, and greatly increased PSD-95 clustering. These results indicate that PKC epsilon promotes synaptogenesis by activating PSD-95 phosphorylation directly through JNK1 and calcium/calmodulin-dependent kinase II and also by inducing expression of PSD-95 and synaptophysin.