Tumor Necrosis Factor-a Converting Enzyme (TACE) Is a Growth Hormone Binding Protein (GHBP) Sheddase: The Metalloprotease TACE/ADAM-17 Is Critical for (PMA-Induced) GH Receptor Proteolysis and GHBP Generation*

The GH binding protein (GHBP), which exists in many vertebrates, is a circulating high affinity binding protein corresponding to the extracellular domain of the GH receptor (GHR). In humans, rabbits, and several other species, the GHBP is generated by proteolysis of the GHR and shedding of its extracellular domain. We previously showed that GHBP shedding is inducible by the phorbol ester phorbol 12myristate,13-acetate (PMA) and inhibited by the metalloprotease inhibitor, Immunex Corp. Compound 3 (IC3). The metzincin metalloprotease, tumor necrosis factor-a (TNF-a)-converting enzyme (TACE), catalyzes the shedding of TNF-a from its transmembrane precursor, a process that is also inhibitable by IC3. TACE may hence be a candidate for GHBP sheddase. In this study, we reconstitute fibroblasts derived from a TACE knockout mouse (Null cells) with either the rabbit (rb) GHR alone (Null/R) or rbGHR plus murine TACE (Null/R1T). Although GHR in both cells was expressed at similar abundance, dimerized normally and caused JAK2 activation in response to GH independent of TACE expression, PMA was unable to generate GHBP from Null/R cells. In contrast, PMA caused ample GHBP generation from TACE reconstituted (Null/R1T) cells, and this GHBP shedding was substantially inhibited by IC3 pretreatment. Corresponding to the induced shedding of GHBP from Null/R1T cells, PMA treatment caused a significant loss of immunoblottable GHR in Null/R1T, but not in Null/R cells. We conclude that TACE is an enzyme required for PMA-induced GHBP shedding and that PMAinduced down-regulation of GHR abundance may in significant measure be attributable to TACE-mediated GHR proteolysis. (Endocrinology 141: 4342–4348, 2000) T GH binding protein (GHBP) corresponds to the circulating extracellular domain of the GH receptor (GHR) (1). It is found in the circulation of many vertebrates and has been evolutionarily conserved from at least the bony fish through humans (2–4). Depending on the species, different mechanisms are used to generate GHBP. In rodents, the GHBP is secreted as an alternative splicing product of the GHR gene that contains a short hydrophilic sequence in place of the transmembrane and intracellular domains (5, 6). This hydrophilic tail is encoded by a special exon (exon 8A) interposed between exons 7 (extracellular) and 8 (transmembrane domain) (7, 8). In many other species, including rabbits and humans, exon 8A is missing, and the GHBP is generated by proteolytic cleavage from the membrane-bound GHR, a process also known as shedding. Although the ultimate physiological role of the GHBP is not known, its evolutionary conservation and the employment of two different mechanisms for its production suggest that it subserves important