AUTOCRINE INDUCTION OF TUMOR PROTEASE PRODUCTION AND INVASION BY A METALLOTHIONEIN-REGULATED TGF-BETA-1 (SER223, 225)

An expression vector was constructed in which TGF-beta-1 was placed under the control of the metallothionein promoter. Cys223 and Cys225 in the TGF-beta-1 propeptide were converted to serines, mutations which result in dissociation of the pro-peptide and secretion of bioactive TGF-beta-1 [Brunner,A.M., Marquardt,H., Malacko,A.R., Lioubin,M.N. and Purchio,A.F. (1989) J. Biol. Chem., 264, 13660-13664]. A fibrosarcoma was transfected with this plasmid and a clone (17.18) was selected in which TGF-beta-1 mRNA was able to be induced six-fold following zinc sulphate treatment. These cells increased the secretion of bioactive TGF-beta-1 14-fold and exhibited a coincidental increase in jun-B mRNA expression, suggesting that secreted TGF-beta-1 was acting to induce this early response gene by autocrine activation. Following zinc sulphate induction, the tumor cells became progressively more motile and able to invade collagen gels. In contrast to parental tumor not bearing the TGF-beta-1 expression vector, zinc sulphate stimulation of clone 17.18 enhanced collagenase IV and procathepsin L mRNA levels and enhanced the secretion of many collagenolytic proteases into the medium. Since the action of TGF-beta generally decreases proteolysis by suppression of protease transcription, we compared the response of normal parental fibroblasts to ras-transformed fibrosarcomas and confirmed that TGF-beta could greatly enhance collagenase IV and procathepsin L mRNA levels while having little effect on non-transformed fibroblasts. These experiments indicate that induction of TGF-beta secretion can enhance motility and protease production through autocrine activation, thus increasing the invasion potential of fibrosarcomas.