Calpain is back: A hypothesis revisited

Calpains are intracellular cystine proteases that are involved several neural processes including long-term potentiation (LTP), learning and memory and synaptic plasticity. In our studies, we demonstrated that m-calpain is activated by MAPK-mediated phosphorylation in neural cells following application of BDNF, EGF or 17-beta Estradiol. We also demonstrated that BDNF and 17-beta Estradiol treatments of neurons resulted in a calpain dependent increase in actin polymerization, specifically in neural dendrites and dendritic spine-like structure. Given the critical role of BDNF in the processes of synaptic plasticity and learning and memory, we also monitored whether BDNF can activate calpain in acute hippocampal slices. Utilizing the DABCYL/EDANS FRET calpain substrate and the CFP/YFP FRET based CAFI mouse model to monitor calpain activation in acute hippocampal slices, our results indicate that BDNF both activates calpain and initiates actin polymerization in the dentate gyrus. Similar results were observed in samples subjected to chemical long term potentiation (cLTP). Both BDNF and cLTP treated samples displayed an increase in AMPA receptors containing GluR1 subunits trafficking as well. Additionally, the functionality of the actin cytoskeletal architecture is dependent on a dynamic equilibrium between polymerized and depolymerized actin. This regulation involves the ADF/cofilin family of proteins. To determine the role of calpain in this actin polymerization machinery, the scaffolding protein 14-3-3 &zgr; was monitored. 14-3-3 &zgr; is a calpain substrate that binds both cofilin and the cofilin phosphastase, Slingshot (SSH). Our results indicate that BDNF and cLTP treatment initiates actin polymerization by inactivating cofilin by phosphorylation. Both phospho-cofilin and phospho-SSH are binding protein, but it is also a substrate of calpain. Association of phospho-cofilin and phospho-SSH with 14-3-3 &zgr;, also sequesters the complex to the cytoplasm following deactivation of cofilin. Our data indicated both BDNF and cLTP treatments resulted in an increase in phospho-SSH and phospho-cofilin association with immunopreciptated 14-3-3 &zgr;. This increase in association was also calpain-dependent suggesting a possible calpain-mediated regulation of 14-3-3 &zgr;, allowing for the formation of such a complex. These results provide a new link between calpain and its participation in cytoskeletal reorganization, synaptic plasticity and learning and memory.