For several years we have engaged in studying the role of two tissue regulators, namely transforming growth factor alpha (TGFalpha) and transforming growth factor beta (TGFbeta) in regulating growth of normal and transformed (RTE) cells. We recently showed that cultures of primary RTE cells secrete small amounts of TGFalpha and two isoforms of TGFbeta, namely TGFbeta1 and TGFbeta2 and that these autocrine growth factors regulate proliferation in the cultures. The role of autocrine TGFbeta in multistage transformation of RTE cells. Upon exposure to physical or chemical carcinogens primary RTE cells undergo a multistep process of neoplastic transformation: The enhanced growth variant colony (EGV), which is the first manifestation of transformation becomes immortalized (=IGV) and subsequently neoplastic (= NGV). We found that late-stage transformants secreted far less TGFB than normal RTE cells (approx. 1/10) and have a greatly reduced responsiveness to the growth inhibitory activity of TGFbeta. Studies with 6-nitrochrysene (6-NC) revealed that this disruption of the TGFbeta system is an early event of RTE transformation. Spontaneous, in contrast to 6-NC induced EGV-colony development was highly sensitive to TGFbeta. A 100-fold higher TGFbeta concentration was required to suppress 6-NC induced EGV-colonies. We concluded that one critical early event in RTE cell transformation is disruption of the TGFbeta system. Characterization of a third TGFbeta1 transcript in cultured RTE cells. We reported that RTE cell cultures express three mRNA transcripts (2.5, 1.9, 1.4, kb). We now show that the 1.9 and 1.4 kb transcripts are not detectable by Northern analysis of resting adult trachea, but are induced in regenerating tracheal epithelium and tumors. Northern analysis of the TGFbeta1 transcripts with subclones of the murine cDNA demonstrates that the 1.4 kb transcript lacks as much of the 5'-untranslated region (UTR). RNase protection analysis was used to map the transcriptional start site of the 1.4 kb transcript to within 100 bp of the first ATG codon. No differences in the coding or 3' UTR were detected between the 1.4 kb and the 2.5 kb transcripts. Because the 1.4 kb transcript is missing most of the long GC-rich 5' UTR, it may be translated at a different rate than the 2.5 and 1.9 kb transcripts, or may code for an intracellular form of TGFbeta1. The 1.4 kb transcript has been observed under several conditions of injury or stress, and therefore may be an important component of the TGFbeta1 response to these conditions.
