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Biochemical Pathways to Long-Term Potentiation

D.D. Gerendasy, J.G. Sutcliffe, A. Dao

RC3/neurogranin plays a role in long-term potentiation, development of dendritic spines, long-term depression, learning, and memory. We proposed that RC3, GAP-43, and the small cerebellum-enriched peptide PEP-19 belong to a protein family that we have named the calpacitins. Membership in this family is based on sequence homology and, we think, a common biochemical function. RC3 and GAP-43 regulate the availability of calmodulin in dendritic spines and axons, respectively, and calmodulin regulates the ability of RC3 and GAP-43 to amplify the mobilization of calcium in response to stimulation of metabotropic glutamate receptors. PEP-19 may serve a similar function in the cerebellum, although biochemical characterization of this molecule has lagged behind that of RC3 and GAP-43.

We suggested that these molecules release calmodulin rapidly in response to large influxes of calcium and slowly in response to small increases. This nonlinear response is analogous to the behavior of a capacitor: hence the name calpacitin. Because calmodulin regulates the ability of RC3 to amplify the effects of metabotropic glutamate receptor agonists, this amplification must have nonlinear kinetics as well. The capacitance of the system is regulated by protein kinase C--mediated phosphorylation of RC3 and GAP-43. Phosphorylated forms of RC3 and GAP-43 do not interact with calmodulin. We further proposed that the ratio of phosphorylated to unphosphorylated RC3 determines the sliding long-term potentiation/long-term depression threshold in concert with calcium/calmodulin-dependent kinase II.

The developmental onset and the cellular and anatomic distributions of RC3 and the neuron-specific isoform of protein kinase C (PKC) are similar, a finding that suggests that this particular isoform phosphorylates RC3. Thus, the phenotypes of PKC knockout mice, which do not have normal long-term potentiation and perform poorly in tasks designed to test spatial memory, could be caused by an inability to phosphorylate RC3. We examined phosphorylation of 3 major neuronal substrates of protein kinase C--RC3, GAP-43, and MARCKS--in these mice after treatment with glutamate or a phorbol ester. Phosphorylation was evaluated with quantitative immunoprecipitation of the 3 protein kinase C substrates from hippocampal slice preparations that had been bathed in medium containing radiolabeled phosphate before treatment. In contrast to the findings with hippocampal slices from control animals, neither phorbol ester nor glutamate stimulated the phosphorylation of RC3 in hippocampal slices from mice that were homozygous for the null mutation. GAP-43 phosphorylation was attenuated in slices from the null mutant, whereas MARCKS phosphorylation was not affected. Our data show that stimulation of RC3 phosphorylation is absent in PKC knockout mouse and defines the following signal transduction pathway: stimulation of the metabotropic glutamate and N-methyl-d-aspartate receptors activate PKC, which in turn phosphorylates RC3. Perturbation of this pathway may be responsible for the physiologic and behavioral phenotypes of PKC knockout mice.

PUBLICATIONS

Gerendasy, D.D., Sutcliffe, J.G. RC3/neurogranin, a postsynaptic calpacitin for setting the response threshold to calcium influxes. Mol. Neurobiol. 15:131, 1997.