GRICH68 AND GRICH70 ARE 2',3'-CYCLIC-NUCLEOTIDE 3'-PHOSPHODIESTERASESINDUCED DURING GOLDFISH OPTIC-NERVE REGENERATION

Biochemical characterization of changes in gene expression that accomp any optic nerve regeneration has led to the identification of proteins that may play key roles in the regeneration process. In this report, a cDNA encoding gRICH70, a novel isoform of the regeneration-induced g RICH68 protein, has been identified and characterized in goldfish. Bot h gRICH68 and gRICH70 show significant homology (34-36%) to mammalian 2',3'-cyclic-nucleotide 3'-phosphodiesterases (CNPases), hence the nam e goldfish regeneration-induced CNPase homolog (gRICH). The predicted 431-amino acid gRICH70 protein is 88% homologous to gRICH68, and the r etinal mRNA for gRICH70 is coordinately induced with gRICH68 mRNA duri ng optic nerve regeneration. Enzymatic analysis of recombinant protein s confirms that both gRICH proteins possess CNPase activity. Despite t he relatively limited sequence homology, the kinetic constants obtaine d suggest that both gRICH proteins are at least as efficient as recomb inant mouse CNP1 in catalyzing the hydrolysis of 2',3'-cAMP. Immunopre cipitation studies indicate that gRICH proteins are responsible for th e majority of the CNPase activity detected in regenerating goldfish re tinas. The evidence presented demonstrates that gRICH68 and gRICH70 co rrespond to a previously described doublet of acidic proteins that are selectively induced in the goldfish retina during optic nerve regener ation, Thus, CNPase enzyme activity is implicated for the first time i n the process of nerve regeneration.