A TRANSFORMING GROWTH-FACTOR-ALPHA PATHWAY IS EXPRESSED IN GH(4)C(1) RAT PITUITARY-TUMORS AND APPEARS NECESSARY FOR TUMOR-FORMATION

Transforming growth factor-alpha (TGF alpha) is a growth regulatory peptide expressed largely as a high mol wt species in the anterior pituitary gland. The overall objective of this work was to test the hypothesis that altered expression of TGF alpha may play a role in the tumorigenicity of the GH(4)C(1) cell line. We examined expression of TGF alpha in three related clones of pituitary tumor cells (GH(1), GH(3), and GH(4)C(1)) grown as transplantable tumors, the MtT/W5 tumor from which they were derived, and anterior pituitary glands of Wistar-Furth rats, the source of the MtT/W5 tumor. Wistar-Furth anterior pituitary, MtT/W5, GH(1), GH(3), and GH(4)C(1) extracts all contained TGF alpha-specific immunoreactivity, which, when examined on sodium dodecyl sulfate-gel transfers, was of high relative mol wt, corresponding to incompletely processed TGF alpha. In neither the anterior pituitary nor the tumors was the fully processed 6-kilodalton TGF alpha form identified, indicating that mature TGF alpha is expressed to only a limited degree in normal and tumor pituitary tissue. We next determined whether a lack of receptors for TGF alpha may account for the MtT/W5 tumor phenotype in vivo. Scatchard analysis of [I-125] epidermal growth factor ([I-125]EGF) saturation isotherm binding identified a comparable class of sites in both the anterior pituitary gland and GH(4)C(1) transplantable tumors. Specific binding sites were also found in MtT/W5, GH(1), and GH(3) tumors. Thus, the functional components of a TGF alpha pathway exist in both the anterior pituitary gland and GH(4)C(1) transplantable tumors. We lastly examined whether a TGF alpha pathway plays a functional role in GH(4)C(1) tumor formation. Toward this aim, we isolated TGF alpha-nonresponsive variants by two different selection schemes: one using a TGF alpha-toxin conjugate, and the other using a TGF alpha-inducible morphological phenotype. Each variant had decreased [I-125]EGF specific binding and little or no EGF growth inhibitory response in vitro. We also isolated a mutagen-induced revertant from one of the variants based on expression of the TGF alpha-inducible morphological phenotype. These cells were found to have a normal complement of receptors and EGF growth inhibitory response in vitro. GH(4)C(1), the two variants, and the revertant cells were inoculated into Wistar-Furth rats, and their growth observed for 8 weeks. The GH(4)C(1) cells and the revertant formed tumors by 8 weeks, whereas the two variant cells failed to form tumors. Taken together, these results indicate that TGF alpha and its receptor are expressed in GH(4)C(1) cells grown in vivo, and that this pathway appears necessary for tumor formation.