The broad long-term goals are to elucidate the molecular events underlying successful neuronal regeneration. Although much is known about regeneration, many molecules required for target innervation remain uncharacterized. The PI has cloned a novel gene, nnal, that is up-regulated in motor neurons following murine sciatic nerve injury. Nnal is down-regulated upon re-innervation of muscle. Nnal is also highly expressed in differentiating nervous system. Sequence analysis of the full length 4kb cDNA identified a nuclear localization signal, an ATP/GTP binding motif and several phosphorylation sites. Based upon motif analysis and expression patterns, it is hypothesized nnal acts as a phosphorylated nuclear ATP/GTP binding protein that regulates events associated with target innervation.
The specific aims will 1) analyze and correlate expression patterns of nnal, 2) test biochemical properties, 3) assess nnal's effect on neurite outgrowth in vitro and 4) determine if nnal is required for successful neuronal regeneration in vivo. The research design uses northern, in situ, western and immunocytochemical analysis to map mRNA and protein expression. Recombinant protein will be used to assess ATP/GTP binding and hydrolysis characteristics. In addition, phosphorylation assays will be used to test the phosphorylation state of nnal in vivo and whether phosphorylation affects nuclear translocation. Finally, adeno-associated viral (AAV) approaches will be used to express mutant nnal in vitro and in vivo. The in vitro assays will examine effects on neurite outgrowth while in vivo assays will assess effects on neuronal regeneration following sciatic nerve injury. In summary, the present application characterizes the role of a completely novel gene, nnal, in regenerative mechanisms. The health relatedness is to understand the molecular cascades required for successful regeneration so that effective treatment strategies can be designed to treat patients suffering from neural injury.
| Ford, Gregory D; Ford, Byron D; Steele Jr, Ernest C et al. (2008) Analysis of transcriptional profiles and functional clustering of global cerebellar gene expression in PCD3J mice. Biochem Biophys Res Commun 377:556-61 |
| Harris, A; Morgan, J I; Pecot, M et al. (2000) Regenerating motor neurons express Nna1, a novel ATP/GTP-binding protein related to zinc carboxypeptidases. Mol Cell Neurosci 16:578-96 |
