Abstract
The Drosophila melanogaster ether-à-go-go (EAG) protein is a voltage gated potassium channel that has a C-terminal tail that binds proteins, such as Ca2+/calmodulin-dependent protein kinase II (CaMKII) and calmodulin (CaM). The interaction between EAG and CaMKII is involved in learning and regulating neuronal excitation (Wang et al., 1994). To study the interaction of EAG and CaMKII, point mutations inserted by homologous recombination into the native eag gene, T787A and F732S/F735S, were investigated in electrophysiological experiments. T787A eliminates the CaMKII phosphorylation target on EAG. F732S/F735S, blocks CaM binding at the C-terminus of \r EAG. Excitatory junction potentials (EJPs) or action potentials were recorded in larval body wall muscles during 1) current injection, 2) fictive crawling motor pattern and 3) nerve stimulation. F732S/F735S was expected to have a decrease in neuron excitability based on a similar mutation in hEAG (Schonherr et al., 2000), while T787A was expected to have an increase in neuron excitability based on oocyte expression (Wang et al., 2002).\r When action potentials were evoked by injecting current, the threshold of one line of F732S/F735S was significantly lower than its control, but the other line had lower, but not significantly lower, threshold. The half decay rate was higher for CaM8 than its three genetic controls, but the genetic control for T787A and F732S/F735S responded differently. The results in the current injection experiment were so far inconclusive. The blocked CaM binding site in EAG might increase neuron excitability and have stronger EAG current. In contrast to the controls, the motor patterns in F732S/F735S CaM mutants were not evoked after separating the ventral nerve cord from larval brain and subesophageal ganglion and adding pilocarpine. The increased excitability across the circuit might lead to increased suppression to compensate particular parts of the circuit. When the nerve was stimulated, there was no significant difference in the shape or number of EJP between the T787A mutant and its control. Electrophysiological studies were used to analyze F732S/F735S and T787A mutants, but more experiments are needed to characterize the interaction between EAG and CaMKII.