A considerable amount of work has been done in recent years on the genetic control of mammalian central nervous system (CNS) development. However, our knowledge of CNS development in general and of rostral CNS specification in particular still remains quite fragmentary.
The proponents set out to adopt a novel methodology based on a recetly devised differential screening technique, to help elucidate some of the molecular mechanisms underlying further subdivision and differentiation of primary brain structures during midgestation brain development.
The aim of the present project is to identify, isolate and analyze novel genes involved at various levels in early brain development, and define their role in normal ontogenesis, and in mutants emerged from the mouse stock or generated by homologous recombination. Our search will not be restricted to genes encoding positional or early patterning information. Instead, the proponents plan to extend our investigation to genes governing various cellular processes. To identify genes displaying restricted, brain-specific or stage-specific expression patterns, the proponents will utilize an updated PCR-based cDNA fingerprinting protocol to analyze tissues obtained from various areas of the brain during early development. The proponents are going to compare patterns obtained from telencephalic vesicles at E10.5 and later stages (temporally restricted expression), as well as patterns from anterior (prosencephalic) and more posterior (mesencephalic) structures at E12.5 (spatially restricted expression). Differentially expressed clones will be sequenced and sequences aligned to online databases through different algorithms (BLAST, FASTA). Six-frame conceptual translations of nucleotide sequences will be analyzed for their coding ability with GRAIL and aligned to protein sequence databases. Multiple alignments and motif searches will be run through the GCG package. Full length clones will be obtained through the screening of custom made mouse embryonic brain libraries; cDNAs will be physically and genetically mapped on the mouse genome; their tissue distribution will be assessed by in situ hybridization of CNS tissue sections. A restricted number of genes of interest will be characterized functionally by a. studies of phenotypic mutants; b. grafting techniques; c. experimental embryology techniques; d. homologous recombination / gene targeting techniques.
Funding SchemeCSC - Cost-sharing contracts