Recombinant DNA clones corresponding to mRNA's induced during successful axon regeneration will be used to evaluate the hypothesis that axon regeneration in many mammalian CNS neurons is limited by failure to induce expression of certain """"""""growth-associated"""""""" genes. Clones representing putative """"""""growth-associated"""""""" genes will be selected by screening a library of clones - containing cDNA complementary to all the mRNA's translated in regenerating toad retinal ganglion cells - for clones which hybridize more readily with radiolabeled cDNA from regenerating retinal ganglion cells (representing total polysomal mRNA) than with the corresponding cDNA from the normal control neurons. Clones selected in this way will be used as probes for the presence of homologous mRNA's in a broad phylogenetic spectrum of neurons during developmental axon growth, successful regeneration, or abortive regeneration. The hypothesis being tested predicts that all neurons undergoing developmental axon growth or successful axon regeneration will show greatly increased abilities to hybridize with the """"""""growth-associated"""""""" cDNA probes (because of increased amounts of """"""""growth-associated"""""""" mRNA in those cells) compared with normal adult neurons, but that mammalian CNS neurons which do not regenerate their axons will not increase their hybridization with the probes after axotomy. Hybridizations of pulse-labeled RNA to the """"""""growth-associated"""""""" DNA clones, cell-free translation of mRNA from normal and regenerating neurons, and gel electrophoresis of in vivo labeled retrogradely transported proteins will be used to investiage mechanisms by which axon interruption leads to specific changes in neuronal gene expression. The proposed research establishes a genetic definition of the differences between neurons which grow axons successfully and those which fail to regenerate injured axons.
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