Transforming growth factor-alpha (TGF alpha), a polypeptide mitogen for cells of epithelial and mesenchymal origin, has been implicated in diverse biological processes including wound healing, cell migration, angiogenesis and bone resorption. A member of the EGF family of growth regulators, it binds to the EGF receptor with high affinity and activates its intrinsic tyrosine kinase. Its precise physiological functions are not known, but TGF alpha is expressed in developing embryos, and in various adult tissues including skin, pituitary gland, decidua and brain, as well as in activated macrophages. The hallmark of TGF alpha expression, however,is the consistent observation that it is most prevalent and abundant in neoplastic cells. Thus, compared to their normal counterparts, the levels of TGF alpha mRNA and protein are elevated in tumors and in cells transformed by chemicals, viruses and activated cellular oncogenes. These data suggest that TGF alpha participates in the development of neoplastic disease and, indeed, TGF alpha promotes the transformation of cells in culture. Additionally, when expressed in transgenic mice it acts as an oncoprotein in mammary gland and liver. Thus, increased expression of TGF alpha appears to be an important event in tumorigenesis though the mechanism by which it is upregulated is unknown. The long term objectives of this proposal are to determine the physiological roles of secreted TGF alpha and its integral-membrane precursor (proTGF alpha), and elucidate the mechanisms that regulate TGF alpha expression in normal and neoplastic cells.
Our specific aims are to: (1) Establish functional roles for TGF alpha by developing mice in which the TGF alpha gene is either overexpressed or globally inactivated. Further characterize lines of transgenic mice that overexpress TGF alpha is either a global or tissue- specific manner. Accomplish the latter via homologous recombination is embryonic stem (ES) cells. Inject correctly targeted ES cells into blastocysts which are then transferred to foster mothers. Establish animals that are chimaeric for inactivation of a TGF alpha allele. Breed these to develop hetero- and homozygous lines. (2) Characterize the biological activities of transmembrane proTGF alpha Derive antibodies to identify cell types in which it might accumulate, and develop lines of transgenic mice that overexpress it globally or in a tissue-specific manner. Use gene targeting to develop mice in which its expression (but not that of the mature, secreted TGF alpha) is abolished. Identify a Drosophila homologue so that its expression, processing and function can be characterized with the aid of fly genetics. (3) Identify mechanisms regulating TGF alpha gene expression in normal and neoplastic cells. Define promoter elements required for constitutive expression, and establish mechanisms by which expression is upregulated in cells transformed by activated oncogenes. Analyze for structural changes that might accompany increased gene expression, and determine the whether changing mRNA stability regulates TGF alpha expression. Finally, establish an endometrial culture system with which to study the mechanism by which estrogen enhances expression of this gene.
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