Understanding the molecular details of neuronal development and regeneration are fundamental issues for both basic and clinical neurobiology. The goal of this project is to understand the molecular mechanisms regulating the spatial and temporal pattern of cytoskeletal rearrangements and their roles in promoting and maintaining neuronal development. The specific aim is to determine the in vivo molecular functions and regulation of microtubule associated proteins (MAPs) during nerve growth factor (NGF) induced microtubule assembly and neurite outgrowth. Our understanding of the role of MAPs in microtubule assembly is derived almost exclusively from in vitro studies with purified microtubule proteins. In order to ascribe a biological relevance to these in vitro studies, a direct demonstration of MAP function in vivo is required. Unfortunately, no direct in vivo experiments have been possible to date. This application aims to use the novel tools and reagents of molecular biology to overcome existing obstacles and directly assess the in vivo roles of MAPs during microtubule assemble and neurite outgrowth. The experiments will examine the role of MAPs during NGF induced neurite outgrowth of PC12 rat pheochromocytoma cells. They will directly assess the recently proposed model in which the microtubule associated proteins MAP1 and tau are limiting during in vivo microtubule assembly and neurite outgrowth in this system and are therefore key regulatory elements controlling both these events. The experimental program involves isolation of a MAP1 cDNA clone using recombinant DNA technology, which will then become the crucial reagent allowing functional and regulatory studies directly testing the above model.