Transforming growth factor alpha: expression, regulation and biological action of its integral membrane precursor

Transforming growth factor alpha (TGF alpha) is a 6 kDa polypeptide mitogen that interacts with the epidermal growth factor receptor and activates its intrinsic tyrosine kinase. The mature 50 amino acid TGF alpha is released from a 159 or 160 amino acid integral membrane glycoprotein precursor, denoted proTGF alpha, via cleavage at both termini by an unknown protease with elastase-like specificity. Rat proTGF alpha is encoded by a 4.5 kb mRNA that is transcribed from a gene containing 6 exons and spanning 85 kb of DNA. Expression of TGF alpha is most prevalent and abundant in transformed cells and tumors, but also detectable at modest levels in a limited number of normal cells and tissues. In many neoplastic cells, proteolytic processing of proTGF alpha is incomplete and/or inefficient, resulting in the preponderance of soluble and/or membrane-bound forms larger than the mature TGF alpha. To characterize the biological activities of the transmembrane TGF alpha precursor in the absence of processing, amino acid substitutions were introduced at the cleavage sites by site-directed mutagenesis of the rat TGF alpha cDNA. Fibroblasts expressing the mutant proTGF alpha constructs did not secrete TGF alpha, but did accumulate proTGF alpha at the cell surface. Coincubation of these cells with A431 cells resulted in binding and autophosphorylation of EGF receptors, and mobilization of intracellular calcium in A431 cells, demonstrating that the transmembrane proTGF alpha can activate EGF receptors on adjacent cells, leading to signal transduction. In addition, rat fibroblasts constitutively expressing the wild-type or mutant proTGF alpha became morphologically transformed in culture, and induced tumors in nude mice. Thus, the interaction between membrane-anchored ligand and receptor triggers mitogenesis that can culminate in neoplastic transformation. To characterize the physiological and pathological effects of TGF alpha in vivo, particularly with respect to epithelial cells, transgenic mice were developed which overexpress the growth factor in multiple or specific tissues. Widespread overexpression of TGF alpha driven by the metallothionein promoter induced epithelial hyperplasia in several organs, including liver and intestine, without disrupting normal tissue architecture. In contrast, the pancreas displayed increased proliferation of both acinar cells and fibroblasts, and focal alteration of acinar cell differentiation. This pancreatic hyperplasia, fibroplasia, and metaplasia were reproduced when TGF alpha expression was placed under control of the elastase promoter, and thus locally restricted to acinar cells, suggesting autocrine and/or paracrine mode of action. Finally, overexpression of TGF alpha promoted neoplastic transformation of certain epithelia. In coagulation gland, there was dramatic hyperplasia and dysplasia with focal evidence of carcinoma in situ.(ABSTRACT TRUNCATED AT 400 WORDS)