Interactions between αCaMKII and calmodulin in living cells: conformational changes arising from CaM -dependent and -independent relationships

Background: alpha CaMKII plays central and essential roles in long-term potentiation (LTP), learning and memory. alpha CaMKII is activated via binding with Ca2+/CaM in response to elevated Ca2+ concentration. Furthermore, prolonged increase in Ca2+ concentration leads to the auto-phosphorylation of alpha CaMKII at T286, maintaining the activation of alpha CaMKII even after Ca2+/CaM dissociation. Importantly, the active form of alpha CaMKII is thought to exhibit conformational change. In order to elucidate the relationships between the interaction of aCaMKII with CaM and the conformational change of alpha CaMKII, we generated molecular probes (YFP-alpha CaMKII with CFP-CaM and YFP-alpha CaMKII-CFP) and performed time-lapse imaging of the interaction with CaM and the conformational change, respectively, in living cells using FRET. Results: The interaction of YFP-alpha CaMKII with CFP-CaM and the conformational change of YFP-alpha CaMKII-CFP were induced simultaneously in response to increased concentrations of Ca2+. Consistent with previous predictions, high levels of Ca2+ signaling maintained the conformational change of YFP-alpha CaMKII-CFP at the time when CFP-CaM was released from YFP-alpha CaMKII. These observations indicated the transfer of alpha CaMKII conformational change from CaM-dependence to CaM-independence. Furthermore, analyses using aCaMKII mutants showed that phosphorylation at T286 and T305/306 played positive and negative roles, respectively, during in vivo interaction with CaM and further suggested that CaM-dependent and CaM-independent conformational changed forms displays similar but distinct structures. Conclusions: Importantly, these structual differences between CaM-dependent and -independent forms of alpha CaMKII may exhibit differential functions for alpha CaMKII, such as interactions with other molecules required for LTP and memory. Our molecular probes could thus be used to identify therapeutic targets for cognitive disorders that are associated with the misregulation of alpha CaMKII.