Screening for mutants that disrupt neurodevelopmental processes in model systems such as Drosophila and the subsequent identification of the causative molecules have been central to understanding of the genetic basis of nervous system development. However, the full promise of forward genetic screening to discern the total complement of genes required for a neurobiological process is rarely realized, in part due to the time and labor required to identify the disrupted genes through conventional genetic mapping techniques. Recent years have seen proof-of-principle studies on the use of Whole Genome Sequencing (WGS) to identify causative point mutations in chemically mutagenized C.elegans or Drosophila strains. In both cases, the strategy was fast and cost-effective. We propose to recruit WGS technology to identify the molecular lesions in a large collection of Drosophila neuromuscular junction (NMJ) synapse mutants we have generated in preliminary studies. Using conventional genetic mapping techniques, we have previously identified the disrupted genes in subset of these mutants and subsequently characterized both novel synaptic regulatory pathways as well mutations in the Drosophila orthologs of human disease relevant proteins. We will determine the causative genetic defect in an additional forty selected synaptic structure mutants with the goal to both increase our understanding of the molecular regulation of synapse development and provide a guide for future, in-depth analysis of the uncovered loci. Furthermore, the repeated, routine application of Whole Genome Sequencing will supply valuable information on the reproducibility and reliability of this approach and establish the technology as a state-of-the-art cloning technique for nervous system mutants in Drosophila and other neurogenetic model systems.
We will conscript the most current high throughput whole genome sequencing and cloning technology and apply it to neurodevelopment mutants. Establishment of this technology as a state-of-the-art method to identify genes that regulate neurodevelopment will rapidly expand the compendium of molecules associated with normal and aberrant nervous system development, many of which are likely to play important roles in human neurological disease.
