| Popis: |
Cotransfection of neo with various oncogenes resulted in CEF transformation in vitro and, in several instances, sarcoma formation in vivo. Transfection of a family of v-src, c-src and v/c-src chimeric constructs demonstrated the ability of the assay to discriminate between transforming and nontransforming genes. Transfection of a number of erbB variants showed that internal mutations, primarily in the kinase domain, contribute significantly to this oncogene's fibroblast transforming abilities. The tumorigenic potentials detected by direct oncogene transfection faithfully reproduced the findings of similar studies using infectious, oncogenic retroviruses. Our studies establish the utility of CEF transformation by direct DNA transfection. Deregulated expression of the TGFα and EGF-R genes has been implicated in the development of a range of mammalian malignancies, most notably those of human origin. While introduction of the TGFα/EGF genes into immortalized rodent cell lines has frequently resulted in cellular transformation, attempts to generate similar results following simple, exogeneous TGFα/EGF treatment have proven largely unsuccessful. The potential role(s) played by these growth factors in the transformation of normal cells, derived from primary cultures has, heretofore, not been closely examined. In chapter 3 it is reported that both normal chicken embryo fibroblasts (CEF) and normal, diploid, human foreskin fibroblasts (HFF) can be efficiently transformed, in an apparent single-step fashion, following exogeneous TGFα/EGF treatment. CEF cells infected with a retrovirus carrying the TGFα gene generated unusually large, aggressively growing soft agar colonies. The ligand-induced transformation of CEF and HFF is affected by culture age. Cultures consistently responded less efficiently as they underwent increasing numbers of population doublings, yet there was no detectable, concomitant, diminution in the number of receptors per cell. This is the first report of efficient transformation of normal cells derived from primary cultures by TGFα and EGF, without the need for other complementing growth factors or oncogenes. |
