Brain-derived neurotrophic factor and epidermal growth factor activate neuronal m-calpain via mitogen-activated protein kinase-dependent phosphorylation.
Calpain is a calcium-dependent protease that plays a significant role in synaptic plasticity, cell motility, and neurodegeneration. Two major calpain isoforms are present in brain, with mu-calpain (calpain1) requiring micromolar calcium concentrations for activation and m-calpain (calpain2) needing millimolar concentrations. Recent studies in fibroblasts indicate that epidermal growth factor (EGF) can activate m-calpain independently of calcium via mitogen-activated protein kinase (MAPK)-mediated phosphorylation. In neurons, MAPK is activated by both brain-derived neurotrophic factor (BDNF) and EGF. We therefore examined whether these growth factors could activate m-calpain by MAPK-dependent phosphorylation using cultured primary neurons and HEK-TrkB cells, both of which express BDNF and EGF receptors. Calpain activation was monitored by quantitative analysis of spectrin degradation and by a fluorescence resonance energy transfer (FRET)-based assay, which assessed the truncation of a calpain-specific peptide flanked by the FRET fluorophore pair DABCYL and EDANS. In both cell types, BDNF and EGF rapidly elicited calpain activation, which was completely blocked by MAPK and calpain inhibitors. BDNF stimulated m-calpain but not mu-calpain serine phosphorylation, an effect also blocked by MAPK inhibitors. Remarkably, BDNF- and EGF-induced calpain activation was preferentially localized in dendrites and dendritic spines of hippocampal neurons and was associated with actin polymerization, which was prevented by calpain inhibition. Our results indicate that, in cultured neurons, both BDNF and EGF activate m-calpain by MAPK-mediated phosphorylation. These results strongly support a role for calpain in synaptic plasticity and may explain why m-calpain, although widely expressed in CNS, requires nonphysiological calcium levels for activation.
Zadran
Sohila
S
Neuroscience Program, University of Southern California, Los Angeles, California 90089-2520, USA.
Jourdi
Hussam
H
Rostamiani
Karoline
K
Qin
Qingyu
Q
Bi
Xiaoning
X
Baudry
Michel
M
D000199:Actins / Q000378:metabolism; D000704:Analysis of Variance; D000818:Animals; D001369:Axons / Q000187:drug effects / Q000378:metabolism; D019208:Brain-Derived Neurotrophic Factor / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D002154:Calpain / Q000378:metabolism*; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D049229:Dendritic Spines / Q000187:drug effects / Q000378:metabolism; D004347:Drug Interactions; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004622:Embryo, Mammalian; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods; D006624:Hippocampus / Q000166:cytology; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009282:Naphthalenesulfonates; D019579:Neocortex / Q000166:cytology; D009474:Neurons / Q000166:cytology / Q000187:drug effects*; D010766:Phosphorylation / Q000187:drug effects / Q000502:physiology; D034741:RNA, Small Interfering / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D020813:Receptor, trkB / Q000235:genetics; D014162:Transfection / Q000379:methods; D004124:p-Dimethylaminoazobenzene / Q000031:analogs & derivatives
D000199:Actins; D019208:Brain-Derived Neurotrophic Factor; D002614:Chelating Agents; D004791:Enzyme Inhibitors; D009282:Naphthalenesulfonates; D034741:RNA, Small Interfering; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; C063126:5-((2-aminoethyl)amino)naphthalene-1-sulfonic acid; D004533:Egtazic Acid; D004815:Epidermal Growth Factor; C063127:4-(4-dimethylaminophenylazo)benzoic acid; D004124:p-Dimethylaminoazobenzene; D020813:Receptor, trkB; D020928:Mitogen-Activated Protein Kinases; D002154:Calpain; C449000:m-calpain; C449002:mu-calpain; D002118:Calcium