A mechanism for Ca2+/calmodulin-dependent protein kinase II clustering at synaptic and nonsynaptic sites based on self-association

The activity of Ca2+/ calmodulin- dependent protein kinase II ( CaMKII) plays an integral role in regulating synaptic development and plasticity. We designed a live- cell- imaging approach to monitor an activity- dependent clustering of green fluorescent protein ( GFP)-CaMKII holoenzymes, termed self- association, a process that we hypothesize contributes to the translocation of CaMKII to synaptic and nonsynaptic sites in activated neurons. We show that GFP- CaMKII self- association in human embryonic kidney 293 ( HEK293) cells requires a catalytic domain and multimeric structure, requires Ca2+ stimulation and a functional Ca2+/ CaM- binding domain, is regulated by cellular pH and Thr286 autophosphorylation, and has variable rates of dissociation depending on Ca2+ levels. Furthermore, we show that the same rules that govern CaMKII self- association in HEK293 cells apply for extrasynaptic and postsynaptic translocation of GFP- CaMKII in hippocampal neurons. Our data support a novel mechanism for targeting CaMKII to postsynaptic sites after neuronal activation. As such, CaMKII may form a scaffold that, in combination with other synaptic proteins, recruits and localizes additional proteins to the postsynaptic density. We discuss the potential function of CaMKII self- association as a tag of synaptic activity.