The objectives of this proposal are to clone the oncogene activated by x-irradiation in a mouse embryo cell line; and from the oncogene construct a probe to investigate oncogene expression. This will be accomplished using transfection and construction of a genomic library. The Lambda vector used will be a Bam H1 substitution vector since it has already been established that Bam H1 does not inactivate the transforming activity of DNA from x-ray transformed cells. The isolated x-ray induced oncogene will then be used to construct a hybridization probe to retrieve the homologous DNA sequences of normal cells (proto-oncogene). DNA sequencing analysis will determine if the oncogene has arisen as a result of mutation of the proto-oncogene. Further, the probe will be used to determine the transcriptional activity of cellular oncogene in normal and x-ray transformed cells. We will then determine possible changes in oncogene activity following x-irradiation. We will also address the hypothesis by regulation of the level of proto-oncogene transcription at the time of irradiation; the transcriptional activity of a gene at the time of irradiation determines the susceptability of that DNA sequence to mutation. Additionally, we will investigate the expression of the proto-oncogene/oncogene when normal when normal and x-ray transformed cells are induced to differentiate. Determining the mechanism of activation of the oncogene by x-irradiation will be important to an understanding of x-ray induced carcinogenesis. The availability and use of a probe for oncogene expression will greatly enhance the knowledge of the mechanism and modulation of oncogenesis by x-rays; and may eventually lead to methods of treatment and prevention of neoplasia in man.
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