Calpain-Cleaved Collapsin Response Mediator Protein-3 Induces Neuronal Death after Glutamate Toxicity and Cerebral Ischemia

Abstract

Collapsin response mediator proteins (CRMPs) mediate growth cone collapse during development, but their roles in adult brains are not clear. Here we report the findings that the full-length CRMP-3 (p63) is a direct target of calpain that cleaves CRMP-3 at the N terminus (ϩ76 amino acid). Interestingly, activated calpain in response to excitotoxicity in vitro and cerebral ischemia in vivo also cleaved CRMP-3, and the cleavage product of CRMP-3 (p54) underwent nuclear translocation during neuronal death. The expression of p54 was colocalized with the terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling-positive nuclei in glutamatetreated cerebellar granule neurons (CGNs) and in ischemic neurons located in the infarct core after focal cerebral ischemia, suggesting that p54 might be involved in neuronal death. Overexpression studies showed that p54, but not p63, caused death of human embryonic kidney cells and CGNs, whereas knock-down CRMP-3 expression by selective small interfering RNA protected neurons against glutamate toxicity. Collectively, these results reveal a novel role of CRMP-3 in that calpain cleavage of CRMP-3 and the subsequent nuclear translocation of the truncated CRMP-3 evokes neuronal death in response to excitotoxicity and cerebral ischemia. Our findings also establish a novel route of how calpain signals neuron death.

Neuroprotection against Traumatic Brain Injury by a Peptide Derived from the Collapsin Response Mediator Protein 2 (CRMP2)

“…The cleavage of CRMP2 by calpain occurs following ischemia (23), neurotrauma (26), excitotoxicity (13), and nerve growth factor deprivation-induced neurite degeneration (24). This cleavage is not specific to CRMP2, however, because CRMP1, -3, -4, and -5 also appear to be cleaved by calpain following neurotoxic insult (23,26,28). In the present study, we demonstrate that a CRMP2 peptide down-regulates dendritic surface expression of NMDARs and protects against neurotoxicity in cell-based assays and in TBI.…”

Phaseic Acid, an Endogenous and Reversible Inhibitor of Glutamate Receptors in Mouse Brain

“…C57B/6 mice (20 -23 g) were obtained from Charles River and bred locally. Under temporary isofluorane anesthesia, MCAO was induced by the intraluminal insertion of a silicon-coated nylon filament (Re L910 PK5, Doccol Corporation) through the common carotid artery into the internal carotid artery and left in place for 60 min as we previously described (20)(21)(22)(23)(24)(25)(26). Cerebral blood flow (CBF) was monitored by laser Doppler flowmetry using a probe located in the ipsilateral parietal bone (1-2 mm posterior to bregma), and a >90% reduction in CSF was considered to indicate successful occlusion.…”

“…The analytes were ionized by negative-ion electrospray using the following conditions: capillary potential 1.75 kV; desolvation gas flow 1100 L/h; cone gas flow 150L/h; and source and desolvation gas temperatures at 120 Cortical neuronal cultures-Primary cortical neurons were prepared from embryonic E15-16 CD1 mice and cultured in neurobasal media supplemented with B-27 and N2 (Invitrogen Canada) for 7 -14 days as previously described (20,24,25). Neurons in these cultures are fully mature and are responsive to glutamate induced excitotoxicity.…”

“…Indirect immunofluorescence staining and confocal microscopy-The procedures for indirect immunofluorescence immunocytochemistry were exactly as described previously (20,22). Mouse monoclonal antibody to PA was a kind gift from Prof E.W.…”

Calpain inhibition prevents amyloid-β-induced neurodegeneration and associated behavioral dysfunction in rats

“…Although the exact mechanism of this process is not fully understood, there is evidence that calpains, Ca 2þ -dependent cystein proteases, are components of the downstream cascade. Inhibition of calpains prevents excitotoxic neuronal cell death in vitro (Caba et al, 2002;Ray et al, 2006) and in vivo (Chiu et al, 2005;Takano et al, 2005), and there is evidence that calpain cleaves several downstream targets that are critical for the progression of excitotoxic neurodegeneration (Hou et al, 2006;Wu et al, 2004). Calpains have therefore been discussed as a target for interference in the neurodegenerative diseases that are associated with neuronal loss (for review see Huang and Wang, 2001;Goll et al, 2003;Zatz and Starling, 2005).…”