Abstract
BackgroundPrion diseases are fatal neurodegenerative disorders that accompany an accumulation of the disease-associated form(s) of prion protein (PrPSc) in the central nervous system. The neuropathological changes in the brain begin with focal deposits of PrPSc, followed by pathomorphological abnormalities of axon terminal degeneration, synaptic loss, atrophy of dendritic trees, and eventual neuronal cell death in the lesions. However, the underlying molecular basis for these neuropathogenic abnormalities is not fully understood.ResultsIn a proteomic analysis of soluble proteins in the brains of mice challenged intracerebrally with scrapie prion (Obihiro I strain), we found that the amount of the full-length form of collapsin response mediator protein-2 (CRMP-2; 61 kDa) decreased in the late stages of the disease, while the amount of its truncated form (56 kDa) increased to comparable levels observed for the full-length form. Detailed analysis by liquid chromatography-electrospray ionization-tandem mass spectrometry showed that the 56-kDa form (named CRMP-2-ΔC) lacked the sequence from serine518 to the C-terminus, including the C-terminal phosphorylation sites important for the regulation of axonal growth and axon-dendrite specification in developing neurons. The invariable size of the mRNA transcript in Northern blot analysis suggested that the truncation was due to post-translational proteolysis. By overexpression of CRMP-2-ΔC in primary cultured neurons, we observed the augmentation of the development of neurite branch tips to the same levels as for CRMP-2T514A/T555A, a non-phosphorylated mimic of the full-length protein. This suggests that the increased level of CRMP-2-ΔC in the brain modulates the integrity of neurons, and may be involved in the pathogenesis of the neuronal abnormalities observed in the late stages of the disease.ConclusionsWe identified the presence of CRMP-2-ΔC in the brain of a murine model of prion disease. Of note, C-terminal truncations of CRMP-2 have been recently observed in models for neurodegenerative disorders such as ischemia, traumatic brain injury, and Wallerian degeneration. While the structural identity of CRMP-2-ΔC in those models remains unknown, the present study should provide clues to the molecular pathology of degenerating neurons in prion diseases in connection with other neurodegenerative disorders.
Highlights
Prion diseases are fatal neurodegenerative disorders that accompany an accumulation of the diseaseassociated form(s) of prion protein (PrPSc) in the central nervous system
The cells positive for green fluorescent protein (GFP)-fluorescence were examined at 4-DIV for neurites longer than 10 μm, and we found that the cells transfected with collapsin response mediator protein-2 (CRMP-2)-ΔC developed 1.2-fold more neurite tips than the cells transfected with the empty vector (Figures 5B and 5C)
Western blot analysis of the brain of mouse [32] and rat [33] showed that CRMP-2A is mainly expressed in the developing brain while CRMP-2B is predominantly expressed in the adult brain
Summary
Prion diseases are fatal neurodegenerative disorders that accompany an accumulation of the diseaseassociated form(s) of prion protein (PrPSc) in the central nervous system. The observation that prnp-/- mice are viable [7] indicates the functional redundancy of PrPC, and raises the question of whether a ‘loss-of-function’ of PrPC is responsible for the neuronal cell death [8]. It is unclear whether PrPSc triggers neuronal cell death [8]. To understand the molecular neuropathology of prion diseases, microarraybased gene expression profiling has been conducted in murine models [9,10,11,12,13] and in cultured cells infected with the scrapie agent [14]. DNA microarray-based approaches [9,10,11,12,13,14] cannot detect the post-translational modifications of proteins, and the proteomic analysis in cultured cells [15] focused on the quantitative changes of proteins rather than their post-translational modifications
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