title: Shedding of the Mer tyrosine kinase receptor is mediated by ADAM17 protein through a pathway involving reactive oxygen species, protein kinase Cδ, and p38 mitogen-activated protein kinase (MAPK).
abstract: Mer tyrosine kinase (MerTK) is an integral membrane protein that is preferentially expressed by phagocytic cells, where it promotes efferocytosis and inhibits inflammatory signaling. Proteolytic cleavage of MerTK at an unidentified site leads to shedding of its soluble ectodomain (soluble MER; sMER), which can inhibit thrombosis in mice and efferocytosis in vitro. Herein, we show that MerTK is cleaved at proline 485 in murine macrophages. Site-directed deletion of 6 amino acids spanning proline 485 rendered MerTK resistant to proteolysis and suppression of efferocytosis by cleavage-inducing stimuli. LPS is a known inducer of MerTK cleavage, and the intracellular signaling pathways required for this action are unknown. LPS/TLR4-mediated generation of sMER required disintegrin and metalloproteinase ADAM17 and was independent of Myd88, instead requiring TRIF adaptor signaling. LPS-induced cleavage was suppressed by deficiency of NADPH oxidase 2 (Nox2) and PKCδ. The addition of the antioxidant N-acetyl cysteine inhibited PKCδ, and silencing of PKCδ inhibited MAPK p38, which was also required. In a mouse model of endotoxemia, we discovered that LPS induced plasma sMER, and this was suppressed by Adam17 deficiency. Thus, a TRIF-mediated pattern recognition receptor signaling cascade requires NADPH oxidase to activate PKCδ and then p38, culminating in ADAM17-mediated proteolysis of MerTK. These findings link innate pattern recognition receptor signaling to proteolytic inactivation of MerTK and generation of sMER and uncover targets to test how MerTK cleavage affects efferocytosis efficiency and inflammation resolution in vivo.
authors.affiliation: Departments of Medicine, Pathology and Cell Biology, and Physiology, and Cellular Biophysics, Columbia University, New York, New York 10032, USA. ebthorp@northwestern.edu
authors.forename: Edward
authors.initials: E
authors.lastname: Thorp
authors.forename: Tomas
authors.initials: T
authors.lastname: Vaisar
authors.forename: Manikandan
authors.initials: M
authors.lastname: Subramanian
authors.forename: Lauren
authors.initials: L
authors.lastname: Mautner
authors.forename: Carl
authors.initials: C
authors.lastname: Blobel
authors.forename: Ira
authors.initials: I
authors.lastname: Tabas
mesh_terms: D051722:ADAM Proteins / Q000737:chemistry / Q000378:metabolism*; D000072198:ADAM17 Protein; D033942:Adaptor Proteins, Vesicular Transport / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D008070:Lipopolysaccharides / Q000494:pharmacology; D013058:Mass Spectrometry; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D053594:Myeloid Differentiation Factor 88 / Q000378:metabolism; D009249:NADP / Q000172:deficiency / Q000378:metabolism; D011392:Proline / Q000378:metabolism; D051745:Protein Kinase C-delta / Q000378:metabolism*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D017384:Sequence Deletion; D015398:Signal Transduction / Q000187:drug effects; D012995:Solubility / Q000187:drug effects; D051197:Toll-Like Receptor 4 / Q000378:metabolism; D000076202:c-Mer Tyrosine Kinase; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*
chemical_list: D051722:ADAM Proteins; D000072198:ADAM17 Protein; D033942:Adaptor Proteins, Vesicular Transport; D008070:Lipopolysaccharides; D053594:Myeloid Differentiation Factor 88; D009249:NADP; D011392:Proline; D051745:Protein Kinase C-delta; D011518:Proto-Oncogene Proteins; D017382:Reactive Oxygen Species; D020794:Receptor Protein-Tyrosine Kinases; D051197:Toll-Like Receptor 4; D000076202:c-Mer Tyrosine Kinase; D048051:p38 Mitogen-Activated Protein Kinases; C476286:Ticam1 protein, mouse; C493487:Tlr4 protein, mouse; C490767:Mertk protein, mouse; C000606463:Adam17 protein, mouse