SITE SPECIFIC INDUCTION OF GENE CONVERSION IN SCHIZOSACCHAROMYCES POMBE

Abstract

We previously showed that alpha-synuclein (-Syn), a protein implicated in the pathogenesis of several neurodegenerative diseases, is a microtubule-associated protein (MAP), facilitating the polymerization of tubulin into microtubules. Therefore, we hypothesized that -Syn might promote neurite outgrowth, a process that requires microtubule assembly. To test this hypothesis, recombinant human wild type (WT) and mutant (A30P and A53T) -Syn proteins were added to cultured primary rat cortical neurons, and their effects on early neurite outgrowth were observed. The WT and mutant -Syn proteins entered the neurons after 1-4 h of incubation. However, a significant increase in neurite outgrowth was observed only in neurons treated with WT -Syn. MES23.5 dopaminergic neuronal cells overexpressing WT -Syn also exhibited enhanced neurite outgrowth, indicating that the ability of -Syn to promote neurite outgrowth was not due to a direct action on the cell membrane or by the membrane translocation process. Co-immunoprecipitation demonstrated that the recombinant human -Syn was bound to tubulin. In addition, the -Syn-treated neurons displayed increased levels of polymerized tubulin. Because -Syn's MAP functionality is mediated by specific domains, we generated N-terminal (a.a. 1-65), non-amyloid- (non-A) component (NAC) (a.a. 61-95) and C-terminal (a.a. 96-140) fragments and added them to the primary neurons. After 1-4 h of incubation, the various -Syn fragments had entered the neurons. However, only the NAC and C-terminal fragments, which have been previously shown to mediate MAP functionality, promoted neurite outgrowth. These results suggest that -Syn promotes neurite outgrowth by facilitating the polymerization of tubulin into microtubules. (PsycINFO Database Record (c) 2013 APA, all rights reserved) (journal abstract)