Shedding of Endogenous Interleukin-6 Receptor (IL-6R) Is Governed by A Disintegrin and Metalloproteinase (ADAM) Proteases while a Full-length IL-6R Isoform Localizes to Circulating Microvesicles.

Generation of the soluble interleukin-6 receptor (sIL-6R) is a prerequisite for pathogenic IL-6 trans-signaling, which constitutes a distinct signaling pathway of the pleiotropic cytokine interleukin-6 (IL-6). Although in vitro experiments using ectopically overexpressed IL-6R and candidate proteases revealed major roles for the metalloproteinases ADAM10 and ADAM17 in IL-6R shedding, the identity of the protease(s) cleaving IL-6R in more physiological settings, or even in vivo, remains unknown. By taking advantage of specific pharmacological inhibitors and primary cells from ADAM-deficient mice we established that endogenous IL-6R of both human and murine origin is shed by ADAM17 in an induced manner, whereas constitutive release of endogenous IL-6R is largely mediated by ADAM10. Although circulating IL-6R levels are altered in various diseases, the origin of blood-borne IL-6R is still poorly understood. It has been shown previously that ADAM17 hypomorphic mice exhibit unaltered levels of serum sIL-6R. Here, by quantification of serum sIL-6R in protease-deficient mice as well as human patients we also excluded ADAM10, ADAM8, neutrophil elastase, cathepsin G, and proteinase 3 from contributing to circulating sIL-6R. Furthermore, we ruled out alternative splicing of the IL-6R mRNA as a potential source of circulating sIL-6R in the mouse. Instead, we found full-length IL-6R on circulating microvesicles, establishing microvesicle release as a novel mechanism for sIL-6R generation.

Senescence-associated release of transmembrane proteins involves proteolytic processing by ADAM17 and microvesicle shedding.

Cellular senescence, a state of persistent cell cycle arrest, has emerged as a potent tumor suppressor mechanism by restricting proliferation of cells at risk for neoplastic transformation. Senescent cells secrete various growth factors, cytokines, and other proteins that can either elicit the clearance of tumor cells or potentially promote tumor progression. In addition, this senescence-associated secretory phenotype (SASP) includes various factors that are synthesized as transmembrane precursors and subsequently converted into their soluble counterparts. Despite the importance of the SASP to tumor biology, it is virtually unknown how transmembrane proteins are released from senescent cancer cells. Here we show in different models of senescence that the metalloprotease A disintegrin and metalloproteinase 17 (ADAM17) is activated and releases the epidermal growth factor receptor ligand amphiregulin and tumor necrosis factor receptor I (TNFRI) from the surface of senescent cells by ectodomain shedding. ADAM17 activation involves phosphorylation of its cytoplasmic tail by mitogen-activated protein kinase (MAPK) p38. Interestingly, unlike amphiregulin and TNFRI, full-length intercellular adhesion molecule 1 (ICAM1) is released from senescent cells by microvesicles independently of ADAM17. Thus, our results suggest that transmembrane proteins can be released by two distinct mechanisms and point to a crucial role for ADAM17 in shaping the secretory profile of senescent cells.