Phosphatidylserine exposure is required for ADAM17 sheddase function

Abstract

ADAM17, a prominent member of the ‘Disintegrin and Metalloproteinase' (ADAM) family, controls vital cellular functions through cleavage of transmembrane substrates. Here we present evidence that surface exposure of phosphatidylserine (PS) is pivotal for ADAM17 to exert sheddase activity. PS exposure is tightly coupled to substrate shedding provoked by diverse ADAM17 activators. PS dependency is demonstrated in the following: (a) in Raji cells undergoing apoptosis; (b) in mutant PSA-3 cells with manipulatable PS content; and (c) in Scott syndrome lymphocytes genetically defunct in their capacity to externalize PS in response to intracellular Ca2+ elevation. Soluble phosphorylserine but not phosphorylcholine inhibits substrate cleavage. The isolated membrane proximal domain (MPD) of ADAM17 binds to PS but not to phosphatidylcholine liposomes. A cationic PS-binding motif is identified in this domain, replacement of which abrogates liposome-binding and renders the protease incapable of cleaving its substrates in cells. We speculate that surface-exposed PS directs the protease to its targets where it then executes its shedding function.

Structural modeling defines transmembrane residues in ADAM17 that are crucial for Rhbdf2/ADAM17-dependent proteolysis

“…It appears likely that different residues in the TMD of ADAM17 will be responsible for regulating the function of Rhbdf1-ADAM17, since replacing the entire TMD of ADAM17 with that of CD62L (also known as L-selectin) or of betacellulin abolished the stimulated shedding of the Rhbdf1 or Rhbdf2 and ADAM17-dependent substrate TGFα without affecting its constitutive shedding (Le Gall et al, 2010). These findings also raise the possibility that the recently reported activation of ADAM17 through phosphatidylserine exposure could, at least in part, regulate ADAM17 through an effect on the interaction of its TMD with that of Rhbdf2 (Sommer et al, 2016). A better understanding of the differential regulation of ADAM17 by Rhbdf1 and Rhbdf2 might aid in the development of selective inhibitors of Rhbdf2-ADAM17, which would block the release of TNFα and of Rhbdf2-selective substrates such as HB-EGF, without affecting the protective function of the Rhbdf1-ADAM17-dependent processing of the substrate TGFα in the skin and intestinal barriers.…”

Proteolytic ectodomain shedding of membrane proteins in mammals—hardware, concepts, and recent developments

“…It is now becoming clear that lipids are not only bystanders, but can directly control proteolytic activity, as is clearly seen for intramembrane sheddases, in particular γ‐secretase and rhomboids (Urban & Wolfe, ; Bondar et al , ; Holmes et al , ; Winkler et al , ). Yet, even the activity of canonical sheddases can be affected by lipids, for example, BACE1 by cholesterol (Ehehalt et al , ; Kalvodova et al , ) or ADAM17 by phosphatidylserine (Sommer et al , ). However, the exact mechanisms by which lipids regulate sheddase activity await further clarification.…”

How membrane asymmetry regulates ADAM17 sheddase function

“…Our investigation originated in the observation that PS-translocation regularly accompanied the activation of the Disintegrin and Metalloproteinase 17 (ADAM17) 3 . This evolutionarily conserved, vitally important protease was originally identified as the TNF-a releasing enzyme.…”