'REGULATORY MECHANISMS OF KALIRIN'S GEF1 DOMAIN'

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DESCRIPTION (provided by applicant): Understanding how neurons initiate neurite outgrowth is important, not just for our basic understanding of development of the nervous system and neuronal morphology, but also to aid in the development of therapies for neurological disorders. My long-term goal is to understand the molecular mechanisms by which guanine nucleotide exchange factors and GTPases modulate neuronal morphology, as a basis for using Kalirin to aid in the development of neuronal regeneration strategies. The goal of this RCDA is to allow me to obtain additional expertise, cultivate several of the skills necessary for an academic scientist, develop a history in a new field of study, and begin to establish a new research program. During the course of this proposal a multifaceted development agenda will involve attending and running journal clubs, giving presentations, taking classes, teaching classes and receiving high-quality mentorship to meet these developmental goals. I will be mentored in multidimensional NMR by Dr. Glenn King, an expert NMR spectroscopist with an outstanding productivity record. Dr. Betty Eipper will help me cultivate the skills needed to manage a new laboratory, teach classes, and train staff. Dr. Eipper has made many seminal contributions to the neuropeptide field and has a long-history of training future leaders in science. Several studies and our preliminary experiments indicate that Kalirin plays a key role in neurite formation and neuronal differentiation. Considering the robust effect of the isolated kalirin GEF1 domain on neuronal development, the goal of this proposal is to better understand the mechanism(s) by which the GEF1 domain of intact kalirin is regulated. We will first identify the domain(s) of kalirin that regulate the GEF1 domain by generating a series of mutant deletion kalirin proteins and testing them in neurite outgrowth and GEF activation assays. Using the information gained from these experiments, we will pursue one of several potential mechanisms by which the identified regulatory domain might control the GEF1 domain. Finally, the mechanistic hypotheses will be further refined at the molecular level by using NMR to determine the three-dimensional structure of the GEF1 with its regulatory elements and examining the effects of ligand interactions upon the structure. Understanding the mechanism by which the GEF domain of kalirin is controlled will provide a basis for future studies to determine the mechanisms by which kalirin controls neuronal morphology.