Nerve Regeneration

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The central nervous system (CNS) of warm-blooded vertebrates does not support nerve regeneration, in contrast to the regenerative potential of the peripheral nervous system (PNS) (1). After injury, CNS axons degenerate resulting in permanent loss of nervous function. This phenomenon has clinical implications for humans and the study of the biochemistry involved in axonal regeneration is of considerable biomedical
interest. In cold-blooded vertebrates the CNS shows marked regenerative potential. The teleost and the amphibian optic nerve have been extensively used as model systems of successful regeneration in the
CNS (2,3). After injury, the axons of the retinal ganglion cells (RGCs) regenerate and reconnect with their targets in the tectum (4-6). Biochemical studies with these systems has led to the identification of several proteins that are induced in neurons that are regenerating their axons, which therefore may play a role in
axonal regeneration. These proteins are known as axonal growth associated proteins (abbreviated as GAPs) and their study is of great importance to understand the regeneration process, the differences of
capacities for regeneration or even for interventions aimed at improving the regeneration response They include cytoskeletal proteins (7,8), cell adhesion proteins (9,10), ion channels (11), transcription factors and other proteins of less well defined function like GAP-43 (12-14) and RICH proteins (7). 

Graduate Student  Gloria Chapa     

 

 
Graduate Student Gloria Chapa

RICH proteins represent a new family of GAPs that was initially shown to be induced in regenerating retinal ganglion cells (RGCs) of goldfish. In goldfish there are two acidic proteins that were designated p68/70 upon their discovery, to reflect their apparent molecular weight (7). The protein doublet was purified from brain tissues and was shown to represent two related proteins partially associated to the plasma membrane (15). The purified proteins were used to generate partial peptide sequences that were used to clone cDNAs encoding p68/70 related proteins (16,17). Sequence analysis showed significant homology to a marker enzyme of mammalian myelin: CNPase (2',3'-cyclic-nucleotide3'-phosphodiesterase) (18). Consequently, the encoded proteins were re-designated gRICH68 and
gRICH70 (for goldfish Regeneration Induced CNPase Homologs of 68 and 70 kDa). The recombinant proteins were expressed both in prokaryotic and eukaryotic systems and it was shown that they possess
2',3'-cyclic-nucleotide 3'-phosphodiesterase activity, identifying the gRICH proteins as novel non-mammalian members of the CNPase family (17).        

A highly specific polyclonal antibody was generated against recombinant gRICH and was used to confirm the identity of the two proteins with the p68/70 doublet components. The antibody was also used in immunodepletion experiments to suggest that these gRICH proteins are the major 2',3'-cyclic-nucleotide 3'-phosphodiesterases in goldfish retinas (17).  Recently, a cDNA encoding a RICH protein has been cloned from a zebrafish library (19). Both the corresponding mRNA and protein (designated zRICH) are induced during regeneration of the optic nerve
in zebrafish. Site directed mutagenesis has identified two residues (H334, T336) in zRICH that are necessary for catalytic activity (19). The zebrafish is emerging as a model system for classic and molecular genetic studies (20,21), offering great potential for future studies aimed at discovering a role of
these proteins in nerve regeneration.