Characterization of novel Drosophila candidate axon guidance molecules

We are dependent on our nervous system functioning correctly for us to move, think, learn, speak and control our bodies. To do this all our nerve cells must connect up in the brain and to the parts of the body they control. Most of this 'wiring together' happens during the growth and development of the embryo in pregnancy. To do this each nerve cell must extend a long process, called an axon, over large distances and through complex environments to find and connect to its appropriate partners. Each axon is guided when to turn and which way to grow to reach its partner by sensing specific chemicals or molecular 'cues' in different parts of the body. These signals are detected by 'receptor' proteins at the tip of the growing axon. We want to find the molecules that work as these receptors or cues to guide axons along their pathways. From what we know already many of the same molecules and receptors in mammals are also present in smaller animals like the fruitfly Drosophila where they do the same job but on a simpler scale. However we also know we haven't found all the molecules needed to wire-up the nervous system. To find additional molecules we have searched through the genomes of mouse, human and Drosophila to find molecules that are similar to one another and to those we already know about which are found in the developing nervous system. We plan to use Drosophila as a model system where we can disrupt or remove the function of these new molecules to find out if and how they act to direct nerve growth. We use Drosophila as a model system so we can rapidly test the role of these new molecules and to allow us to reduce the need to sacrifice large numbers of mice in research. Once we have found out how these molecules work in Drosophila we will inform other researchers so that the molecules can be tested in other model systems. We need this information both to learn how the nervous system is made and to find out what molecules might be useful in helping us to repair neural injuries or diseases that lead to paralysis or neural degeneration. Unfortunately mammals cannot repair nerve damage that occurs in the brain, our hope is that by identifying the molecules that were originally used to drive and direct nerve cell growth in the embryo we can re-supply these molecules to help nerve cell regeneration in people.

The formation of a correctly functioning nervous system requires a large number of neurons to establish a precise pattern of inter-connectivity. Each neuron extends an axonal projection along a stereotypical path from its cell body to its final target. Both vertebrates and invertebrates face the same formidable task of establishing an intricate pattern of axon pathways during development. It is clear from the molecules that have been identified thusfar with a role in this process that many are highly conserved. However the numbers of molecules identified to date are likely to be insufficient to encode the complete nervous system wiring. We have completed a comparative screen of the mouse, human and Drosophila genomes to define a set of conserved novel transmembrane proteins that possess novel combinations of structural domains previously identified within known axon guidance molecules. Of these a small group of previously uncharacterized Drosophila molecules are expressed specifically within the CNS during axon extension. These molecules can be placed into several classes (i) those that have specific homologues in mouse and human (ii) those that have multiple related vertebrate molecules and (iii) those that appear to be specific to Drosophila. We plan to investigate the precise expression of these molecules and their functional requirement during neural development in vivo by removing their function in the embryo through gene knockout or RNA interference. We will investigate the requirement for the individual domains through the use of gain-of-function and rescue assays. Through this analysis we hope to identify the structural domain combinations that have conserved, widespread or specific roles in directing axonal growth. We anticipate that this work will lead to the identification of novel molecules or motif combinations that have a direct role in axon guidance and may lead to the characterization of reagents to aid neural regeneration.