CLONING OF GENES INVOLVED IN CELL IMMORTALIZATION

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DESCRIPTION: Dr. McCormick is studying the nature of the changes causally involved in transforming normal human cells into cancer cells. Strong evidence indicates that escape from senescence is one of the fundamental changes involved. They have succeeded in generating two infinite life span human fibroblast cell strains, MSU-1.0 and MSU-2, which arose from clonally-derived populations transfected with a v-myc gene. They and others have data indicating that upregulation of expression or deregulation of expression of the myc gene contributes to immortalization of human mesenchymal cells, but is not sufficient to allow them to escape from senescence. Cells must also lose the activity of both alleles of a gene(s) that prevents such escape. The aim of this proposal is to identify and characterize such myc-complementing gene or genes whose activity must be lost for such cells to become immortal. They have been examining this question using mRNA Differential Display (DD) for the diploid strain MSU-1.0 and have found that there are a significant number of down-regulated genes compared with the level of expression in their parental LG1 cells. They will test the candidate genes so identified for their ability to return the MSU 1.0 cells to the mortal state. If these studies are not successful in identifying the gene(s) of interest, they will examine the genes that are up-regulated in these cells. Although the usual interpretation of the data of cell fusion studies is that immortalization involves down-regulation of genes, one can build models in which expression of genes are up-regulated, e.g. the telomerase gene. If they do not succeed in identifying the myc-complementing gene in the studies with MSU 1.0 cells, they will carry out DD on two other immortal human fibroblast cell lines (MSU-2 and OUMS-24F) and their respective isogeneic mortal parental cells (SL80 and OUMS). Human sarcomas, as well as human sarcoma-derived cell lines, frequently exhibit up-regulation of myc. Thirty human sarcoma cell lines are available in their laboratory. To increase the chances of success in identifying the gene(s) of interest, they will include in their analysis of the MSU-2 and OUMS-24F cells, an analysis of the levels of expression of genes found in those human sarcoma derived cell lines that they identify as overexpressing myc. Candidate genes identified by DD will be tested for ability to return the cells to the mortal state.