The overall objective is to understand the mechanisms by which activated proto-oncogenes alter normal growth regulatory processes. This will be accomplished by the use of both molecular (i.e. Northern hybridizations and nuclear run-off assays) and cell biological technique (i.e. growth factor requirements for G1 traverse and logarithmic growth. C3H/10t1/2 cells are not stimulated to grow in soft agar by addition of TGFBeta. We have obtained preliminary data demonstrating that transfection of C3H/10T1/2 cells with a mouse c-myc gene linked to an SV40 promoter results in responsiveness to TGFBeta as assayed by colony formation in soft agar. In addition, transfection with an activated H-ras gene results in a transformed cellular morphology, spontaneous growth in soft agar and markedly decreased binding of 125I-labeled TGFBeta. It is proposed that transfection with the activated H-ras gene abrogated proliferative restraints by inappropriate transcriptional and translational activity in growth factor or cell cycle regulated genes normally utilized during controlled proliferation. The myc gene, however, is suggested to be a modulator, rather than inducer of various growth promoting signals. Thus, an optimal (not necessarily appropriate) growth response will result from the synergistic action of both proto-oncogenes. These hypotheses will be addressed by the following specific aims: 1) Determining whether C3H/10T1/2 cells transfected with an activated H-ras and/or myc gene show transcriptional activity of genes known to be under growth factor control; 2) Determining the effects of ras and/or myc gene transfection in C3H/10T1/2 cells on the in vitro production of specific growth factors; 3) Determining the growth factor requirements for G1 traverse and logarithmic growth of ras and/or myc gene transfected cells; and 4) Further characterization of the growth factor specificity for anchorage-independent growth of myc gene transfected C3H/10T1/2 cells. Overall, these studies should increase our knowledge of the mechanisms controlling normal cellular proliferation as well as transformation.
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