Recombinant type 1 transforming growth factor beta (TGF-,) was expressed to high levels in CHO cells by using dihydrofolate reductase (dhfr) gene amplification. The expression plasmid was derived from the pSV2 vectors and contained, in tandem, the simian TGF-I and mouse dhfr cDNAs. Transcription of both cDNAs was controlled by the simian virus 40 early promoter. Stepwise selection of transfected CHO cells in increasing concentrations of methotrexate yielded cell lines that expressed amplified TGF-, nucleic acid sequences. The expression plasmid DNA was amplified greater than 35-fold in one of the methotrexate-selected transfectants.The major proteins secreted by these cells consisted of latent as judged by immunoblots by using site-specific anti-peptide antibodies derived from various regions of the TGF-, precursor. Levels of recombinant TGF-0 protein secreted by these cells approached 30 ,ug/24 h per 107 cells and required prior acidification for optimal activity; nonacidified supernatants were approximately 1% as active as acidified material. Antibodies directed toward sequences present in the mature growth factor readily identified a proteolytically processed recombinant TGF-, which, on sodium dodecyl sulfate-polyacrylamide gels, comigrated with highly purified natural TGF-4. In addition to mature recombinant TGF-jA, site-specific antibodies demonstrated the existence of larger TGF-3 precursor polypeptides. The availability of biologically active recombinant type 1 TGF-I8 and precursor forms should provide a means to examine the structure, function, and potential in vivo therapeutic use of this growth factor.Results of recent studies have suggested that transforming growth factor beta (TGF-P) consists of a family of closely related biologically active polypeptides with potent cellularmodulating activities. Three molecular forms of TGF-P (types 1, 2 and 1.2) have been identified and characterized.Type 1 TGF-P was the first growth factor isolated and, as a result, the best characterized (for reviews, see references 23 and 47). sources have been obtained. The predicted protein sequences derived from the cDNAs reveal a remarkable homology. The mature forms of the human and simian growth factor differ from the murine factor by one amino acid. Analysis of the various TGF-,B cDNA clones suggests that the mature 112 amino acid chain of type 1 TGF-P is derived from a much larger precursor polypeptide by proteolytic cleavage. The full-length precursor of TGF-,1 shows a high degree of structural conservation and constitutes 391 amino acids for human and 390 amino acids for both murine and simian precursors. A typical leader sequence is observed in the precursor, as well as three potential N-linked glycosylation sites. A dibasic sequence immediately precedes the amino-terminal alanine residue of the mature growth factor, indicating the involvement of a dibasic protease in processing of the growth factor.To understand in more detail the structure and processing of type 1 TGF-P, as well as to provide a source for large...

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