The goal of this project is to dissect the genes involved in the signal transduction pathways of TGF-B leading to inhibition of growth. Since insertional mutagenesis with a retrovirus is not an efficient way to isolate mutant cells, the chemical mutagen, nitrosoguanidine, was used to mutagenize mink lung cells. Mutant cells which have been isolated are resistant to inhibition of growth by TGF-B. Work has begun on construction of CDNA libraries from modified CDM-8 cells which are sensitive to inhibition by TGF-B. This will facilitate the isolation of revertants which are sensitive to inhibition by TGF-B by using a BrdU-Hoechest-visible light treatment. These cell lines will be used to identify genes involved in signal transduction by TGF-B. Another way to identify genes involved in signal transduction is also being developed. Since mutagenized cells are mixtures of TGF-B sensitive and resistant cells, the BrdU-Hoechest-visible light selection method was used to eliminate cells which are resistant to inhibition by TGF-B. The cells which are sensitive to growth inhibition by TGF-B are being treated with the retrovirus containing the suppressor gene from E. coli by cloning in the lambda vector. These studies will increase our understanding of the mechanisms of signal transduction of TGF-B leading to growth inhibition.
