Chromosome abnormalities including aneuploidy, deletions, amplifications, translocations, and other rearrangements are the hall-mark of neoplasia. Until recently the causal role of these genomic changes was questioned, but the avalanche of new findings in this field has made it evident that they play a fundamental role in neoplasia. The long-range goals of ongoing research and of the new projects outlined in this proposal are (i) to elucidate the central role of chromosome instability in the origin of neoplasia, especially to investigate its molecular basis and (ii) to identify and clone genes that suppress tumor formation; we postulate that rearrangements can inactivate suppressor genes. The special system that I developed, a unique Chinese hamster fibroblastic cell line (CHEF) has made possible an integrated genetic, cytogenetic, and molecular analysis now in mid-stream. Many of the results obtained in our ongoing research provide leads into future research, including (1) specific chromosome aberrations in tumor derived CHEF cells; (2) """"""""hit and run"""""""" tumorigenesis; (3) amplification studies showing that sites are localized in transformed cells but dispersed in normal cells; (4) chromosome rearrangements induced in diploid CHEF/16 cells during tumor formation but not during growth in culture; (5) methylation changes in tumor-derived compared with normal CHEF cells; (6) discovery of a site specific endonuclease that nicks dGdT stretches in Z-DNA; (7) demonstration of the resistance of normal human cells (but not CHEF cells) to transformation by oncogene DNA that was introduced by transfection and is being expressed; (8) suppression of tumor formation in hybrids from fusions of normal x tumor CHEF cells, including oncogene-transfected CHEF cells. The major themes of projected research stress molecular approaches based on prior cellular evidence. Evidence of transposition as the molecular basis of instability will be sought inter alia by methods derived from successful work with other organisms. Selection of unique cDNAs by differential or subtractive hybridization, as well as selection of suppressed transfectants, will be used to isolate tumor suppressor genes.
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