title: Neutrophil elastase acts as a biased agonist for proteinase-activated receptor-2 (PAR2).
abstract: Human neutrophil proteinases (elastase, proteinase-3, and cathepsin-G) are released at sites of acute inflammation. We hypothesized that these inflammation-associated proteinases can affect cell signaling by targeting proteinase-activated receptor-2 (PAR(2)). The PAR family of G protein-coupled receptors is triggered by a unique mechanism involving the proteolytic unmasking of an N-terminal self-activating tethered ligand (TL). Proteinases can either activate PAR signaling by unmasking the TL sequence or disarm the receptor for subsequent enzyme activation by cleaving downstream from the TL sequence. We found that none of neutrophil elastase, cathepsin-G, and proteinase-3 can activate G(q)-coupled PAR(2) calcium signaling; but all of these proteinases can disarm PAR(2), releasing the N-terminal TL sequence, thereby preventing G(q)-coupled PAR(2) signaling by trypsin. Interestingly, elastase (but neither cathepsin-G nor proteinase-3) causes a TL-independent PAR(2)-mediated activation of MAPK that, unlike the canonical trypsin activation, does not involve either receptor internalization or recruitment of β-arrestin. Cleavage of synthetic peptides derived from the extracellular N terminus of PAR(2), downstream of the TL sequence, demonstrated distinct proteolytic sites for all three neutrophil-derived enzymes. We conclude that in inflammation, neutrophil proteinases can modulate PAR(2) signaling by preventing/disarming the G(q)/calcium signal pathway and, via elastase, can selectively activate the p44/42 MAPK pathway. Our data illustrate a new mode of PAR regulation that involves biased PAR(2) signaling by neutrophil elastase and a disarming/silencing effect of cathepsin-G and proteinase-3.
authors.affiliation: Department of Physiology and Pharmacology, University of Calgary, Calgary, Alberta T2N 4N1, Canada. rramacha@ucalgary.ca
authors.forename: Rithwik
authors.initials: R
authors.lastname: Ramachandran
authors.forename: Koichiro
authors.initials: K
authors.lastname: Mihara
authors.forename: Hyunjae
authors.initials: H
authors.lastname: Chung
authors.forename: Bernard
authors.initials: B
authors.lastname: Renaux
authors.forename: Chang S
authors.initials: CS
authors.lastname: Lau
authors.forename: Daniel A
authors.initials: DA
authors.lastname: Muruve
authors.forename: Kathryn A
authors.initials: KA
authors.lastname: DeFea
authors.forename: Michel
authors.initials: M
authors.lastname: Bouvier
authors.forename: Morley D
authors.initials: MD
authors.lastname: Hollenberg
mesh_terms: D000818:Animals; D019390:Arrestins / Q000235:genetics / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D056649:Cathepsin G / Q000235:genetics / Q000378:metabolism; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007249:Inflammation / Q000235:genetics / Q000378:metabolism; D019272:Leukocyte Elastase; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000235:genetics / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000235:genetics / Q000378:metabolism; D053146:Myeloblastin / Q000235:genetics / Q000378:metabolism; D010455:Peptides / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D051381:Rats; D044464:Receptor, PAR-2 / Q000819:agonists* / Q000235:genetics / Q000378:metabolism*; D000071557:beta-Arrestins
chemical_list: D019390:Arrestins; D010455:Peptides; D044464:Receptor, PAR-2; D000071557:beta-Arrestins; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; C534094:CTSG protein, human; D056649:Cathepsin G; C534097:Ctsg protein, rat; D019272:Leukocyte Elastase; D053146:Myeloblastin; D043802:GTP-Binding Protein alpha Subunits, Gq-G11