The alpha-fetoprotein (AFP) gene is of special importance because it is an embryonic and fetal gene that is inactivated during development, repressed by corticosteroids, and reactivated by carcinogenesis. The proposed research will define transcription controls of the AFP gene and determine how neoplasia alters them. Recombinant DNA constructs in which AFP gene elements were fused to the bacterial """"""""CAT"""""""" gene have been developed, defining upstream enhancer and modulatory elements that allow expression in the human hepatoma cell line, HepG2. One group of experiments will be directed to better defining the transcription and hormonal controls, by sequencing the DNA of the transcription control region, resolving the control sequences by deletion analysis, and analyzing the transcription- control effects in HepG2 cells. These cell systems will be modified to allow evaluation of hormonal interactions with the AFP-gene control-elements. A second group of studies is directed to the analysis of AFP gene controls in other cells, which normally express AFP, but seem to require more genetic information than is contained in the plasmids already tested. Such cells probably reflect gene controls more like those required for normal development. For this phase of analysis, more DNA regions flanking the AFP gene will be cloned and analyzed. A final group of experiments will study the modifications of cellular AFP gene expression and gene structure caused by the presence in cells of specific activated oncogenes. These experiments will be carried out in preexisting tumor-cell lines and in new cell lines derived by transformation of primary hepatocyte cultures.
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