The overall goal of this Phase I proposal is to discover full rosters of genes involved in signal transduction and oncogenesis. We will concentrate on human GTP binding protein (G-protein) genes and G- protein coupled receptor (GCR) genes, only a fraction of which are known. Discovery of these novel genes will then permit studies of their role in neoplastic transformation. As members of gene families sharing functional characteristics, G protein and GCRs are amenable to discovery by """"""""functional probing"""""""" of genomic libraries. We will derive degenerate DNA probes (oligonucleotides, PCR amplicons) from highly conserved protein motifs of thromboxane receptors, bombesin receptors, thrombin receptors and Gq alpha proteins. We will optimize conditions for hybridization of these probes to gridded human genomic DNA libraries of large-insert (80-150 kb) PI and PAC (PI Artificial Chromosome) clones. The positive genomic clones will be confirmed by Southern analysis with the motif probe and characterized by cytogenetic mapping and genomic sequencing. A unique identifier tag will be derived for each novel gene family member and used to assess expression of different transcripts in normal tissues and in cell lines derived from small cell lung and medullary thyroid cancers.
Genomic clones will be assembled into gene family kits and sold to researchers in basic and applied pharmaceutical research. In addition, sequences specific to individual family members will be also commercialized as probes and primers for gene expression analysis. Commercial value of these resources is high, as they constitute targets for drug discovery and resources for basic research on signal transduction derangements in cancer.
