Functional Study Of Calmodulin Acetylation

Lysine acetylation plays an important role in the celluar functions,which is dynamic and reversible.Most of the researches are mainly focus on the histone acetylation,which can regulate the structure of chromatin and promote transcription.With the development of mass spectrum technology,more and more studies show that there are many non-histone proteins which can be acetylated outside the cell nucleus.In the central nervous system,histone acetylation plays a vital role in a variety of physiological processes,such as synaptic development and plasticity,learning and memory and cognition.However,very little is known about the function of acetylation of non-histone proteins in the central nervous system.Our previous studies identified more than one thousthand acetylated proteins which included calmodulin.Calmodulin(CaM)is Ca2+-binding protein which can bind with many target proteins and activate downstream celluar signaling pathway.CaM is one of the most abundant proteins in postsynaptic density.It can bind with and activate CaM-dependent kinase??(CaMKII?)when NMDA receptor-dependent LTP(long-term potentiation)occurs.Then the activated CaMKIIa triggers a phosphorylation cascade.This signaling pathway is one of the most important mechanisms in regulating LTP.However,whether CaM acetylation has effects on NMDA receptor-Ca2+-CaM-CaMKII?pathway was not investigated.In this study,we performed a series of experiments,such as cell culture,primary neuron culture,chemical-induced LTP,plasmid construction,protein expression and purification,western blotting,immunoprecipitation and protein-protein interaction.Our main results were as follows:first,CaM could be acetylated in 293T cell probably by acetyltransferase CBP,second,chemical LTP could increase CaM acetylation through an NMDA receptor-dependent manner,lastly,acetylation mimicking mutant of CaM has higher affinity with Ca2+ and is more effective to activate CaMKII? when compared with wild type CaM.In sum,these results revealed important roles of CaM acetylation in regulating NMDA receptor-Ca2+-CaM-CaMKIIa pathway,which was necessary for the future study of CaM acetylation influence on synaptic plasticity such as LTP.