The studies of the molecular mechanism of inherited susceptibility to cancer are significantly aided by the identification and analysis of predisposing genes from familial cancer syndromes such as retinoblastoma, Wilms'Tumors, Neurofibromatosis Type I, and Familial Polyposis Coli. The tumor suppressor gene p53 has been shown to frequently altered in a wide variety of sporatic tumors. In these instances, one allele of p53 has been lost and the remaining allele has acquired a somatic mutation. The identification of a germ-line p53 mutation in cancer-prone families with Li-Fraumeni syndrome (LFS) has introduced a new role for p53, i.e., a role in genetic susceptibility to cancer. Our recent studies show that the normal skin fibroblast (NSFs) cells derived from individuals in the cancer- prone family express low levels of both wild-type (wt) and mutant (mt) p 53 protein at levels comparable to that of the wt p53 detected in normal cells. Furthermore, tumors arising from two germ-layers in four different individuals in this family exhibited the loss of the wt p53 allele and the retention of the mt allele. We have also known that many of the mutations in p53 found in LFS families appear to have a transdominant effect over the wt p53 in in vitro assays. To understand the role of germ-line p53 mutations in genetic predisposition and tumorigenesis, we propose to analyze the biological and biochemical characteristics of mt p53 found in this cancer-prone family with LFS. Therefore, we will examine the SV40 T Ag, MDM-2 and sequence specific DNA binding properties of p53 in these NSFs, and evaluate the effect of this wt/mt phenotype on cell cycle progression and apoptosis. Such analysis will provide information on the effects of mt p53 on the functions of wt p53 in a heterozygous situation. We will extend these studies to other LFS families in order to understand the broad implications of these findings for inherited p53 mutations. Since the p53 protein exhibits the properties of a transcription factor and binds to specific DNA sequences in vitro, the family NSFs with endogenous expression of both mt and wt p53 provide a good experimental system with which to identify target genes modulated by mutation in p53. The status of such genes will be evaluated by the analysis of either the expression of previously identified cell growth associated genes or the expression of genes which are differentially regulated in family NSFs (wt/mt). Thus, the experiments proposed in this application should serve not only to increase our understanding of the functional implications of mt p53 in predisposition, tumor formation and progression in cancer-prone families with LFS but should also yield information relevant to the function of p53 in tumorigenesis in general.
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