Calpain‐mediated cleavage of collapsin response mediator protein(CRMP)‐2 during neurite degeneration in mice

Abstract

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.

Vitamin E Deficiency Induces Axonal Degeneration in Mouse Hippocampal Neurons

“…We measured collapsin response mediator protein (CRMP)-2 to determine the mechanisms of morphological change in hippocampal neurons. CRMP-2, which is part of a family of cytoplasmic proteins in the brain, plays important roles in axon guidance (14,18,19). In this study, we found axonal degeneration and a change in CRMP-2 expression in the hippocampal neurons of vitamin E-deficient mice.…”

“…Axonal degeneration was evaluated by monitoring morphological hallmarks of neurite degeneration such as beading, accumulation and fragmentation, as described previously with some modifications (14,21,22). Photomicrographs of the hippocampal CA1 region on the Bodian's stained slices were analyzed to determine axonal condition.…”

“…CRMP-2 plays a pivotal role in neurite functions (14,18,19), binding to tubulin and connecting with the microtubule assembly. Changes in CRMP-2 may be expected to influence the axonal transport system.…”

“…To address this problem we focused on the axons and dendrites, as they play an important role in neurotransmission (13,14). In the axonal transport system, axons are involved in carrying several substances including neurotransmitters and other proteins.…”

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

“…Later studies identified that the cleavage was by calpain (13,(22)(23)(24)(25)(26), a Ca 2ϩ -activated protease that has been heavily linked to neurotoxic signaling (27). The cleavage of CRMP2 by calpain occurs following ischemia (23), neurotrauma (26), excitotoxicity (13), and nerve growth factor deprivation-induced neurite degeneration (24).…”