Abstract
ADAM17, prominent member of the "Disintegrin and Metalloproteinase" (ADAM) family, controls vital cellular functions through cleavage of transmembrane substrates. Several of these play central roles in oncogenesis and inflammation, yet despite its importance, the mechanism by which ADAM17 is activated is not fully understood. We recently presented evidence that surface exposure of phosphatidylserine (PS) is the penultimate event required for sheddase activation, which occurs upon binding of a membrane-proximal, cationic binding motif to the anionic phospholipid headgroup. Here, we show that mutagenesis of the 3 amino acids constituting the PS-binding motif leads to embryonic lethality in mice. Heterozygotes showed no abnormalities. Primary hepatocytes and fibroblasts were analysed and found to express the mutant protease on the cell surface. However, PMA-stimulated release of ADAM17 substrates was completely abolished. The results directly support the novel concept of transiently externalised PS as essential trigger of extracellular protease function in vivo.
Highlights
ADAM17, originally discovered as the TNF-α converting enzyme [1, 2, 3], has emerged as a pre-eminent member of the “a disintegrin and metalloprotease” family of transmembrane proteinases
The starting point of our investigation was the observation that sheddase function of ADAM17 apparently required interaction of the protease with surface-exposed PS [17]. 3D-heteronuclear nuclear magnetic resonance experiments identified a small cluster of cationic amino acids R625/K626/G627/ K628 as the putative PS-binding motif
We provide in vivo evidence in support of the contention that surface-exposed PS plays a critical role in the control of ADAM17 sheddase function
Summary
ADAM17, originally discovered as the TNF-α converting enzyme [1, 2, 3], has emerged as a pre-eminent member of the “a disintegrin and metalloprotease” family of transmembrane proteinases. More than 80 ADAM17 targets have been identified to date, prominent among which are cytokines, adhesion molecules, and cell surface receptors, including TNFR1 [4, 5, 6, 7]. ADAM17 regulates cell growth through the liberation of epidermal growth factor receptor (EGFR) ligands such as amphiregulin (AREG) or TGF-α [8, 9, 10]. ADAM17 deficiency results in severe inflammatory skin and bowel disease, underlining its important role for epithelial cell homeostasis [11, 12]. In vivo mouse models further emphasise the vital importance of ADAM17 [3, 13, 14, 15, 16, 17]. Targeted deletion of exon 11 encoding the catalytic site of the protease (TaceΔZn/ΔZn) [3] resulted in the death of mice between embryonic day (E) 17 and the first day after birth because of developmental defects in brain, heart, lung, skin, skeletal, and immune system [3, 18]
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