Calpain-mediated cleavage of collapsin response mediator protein(CRMP)-2 during neurite degeneration in mice.
Axon or dendrite degeneration involves activation of the ubiquitin-proteasome system, failure to maintain neuritic ATP levels, microtubule fragmentation and a mitochondrial permeability transition that occur independently of the somal death programs. To gain further insight into the neurite degeneration mechanims we have compared two-dimensional gel electrophoresis patterns of neurite proteins from suprior cervical ganglia during degeneration caused by nerve growth factor (NGF) deprivation. We show here that collapsin response mediator protein (CRMP)-2 and CMRP-4 protein patterns were altered during beading formation, an early hallmark of neurite degeneration, prior to neurite fragmentation, the final stage of degeneration. Western blotting using a monoclonal antibody against CRMP-2 shows that the native form (64 kDa) was cleaved to generate a truncated form (58 kDa). No cleavage of CRMP-2 or -4 occurred in NGF-deprived neurites from Wld(s) (Wallerian degeneration slow) mutant mice in which neurite degeneration is markedly delayed. Using different protease inhibitors, purified calpain 1 protein and calpain 1-specific siRNA, we have demonstrated that CRMP-2 is a substrate for calpain 1. Indeed, caplain activity was activated at an early phase of neuronal degeneration in cerebellar granule neurons, and down-regulation of caplain 1 expression suppressed CRMP-2 cleavage. Furthermore, this cleavage occurred after vinblastine treatment or in vitro Wallerian degeneration, suggesting that it represents a common step in the process of dying neurites. CRMP-2 and -4 play a pivotal role in axonal growth and transport, and the C-terminus region of CRMP-2 is essential for its binding to kinesin-1. Hence, this cleavage will render them dysfunctional and subject to autophagic processing associated with beading formation, as evidenced by the finding that the truncated form was localized in the beadings.
Touma
Ekatherina
E
Molecular Neurobiology Laboratory, Graduate School of Life Science, Hokkaido University, Sapporo 060-0810, Japan.
Kato
Satoko
S
Fukui
Koji
K
Koike
Tatsuro
T
D000818:Animals; D002154:Calpain / Q000378:metabolism*; D002478:Cells, Cultured; D002531:Cerebellum / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology; D015180:Electrophoresis, Gel, Two-Dimensional; D004789:Enzyme Activation; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D009410:Nerve Degeneration / Q000209:etiology / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D020932:Nerve Growth Factor / Q000172:deficiency; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D016501:Neurites / Q000378:metabolism*; D010446:Peptide Fragments / Q000096:biosynthesis; D017783:Superior Cervical Ganglion / Q000378:metabolism / Q000648:ultrastructure; D014855:Wallerian Degeneration / Q000378:metabolism
C526904:Dpysl3 protein, mouse; D036341:Intercellular Signaling Peptides and Proteins; D009419:Nerve Tissue Proteins; D010446:Peptide Fragments; C095114:collapsin response mediator protein-2; D020932:Nerve Growth Factor; D002154:Calpain; C112682:Capn1 protein, mouse