title: Mechanisms of desensitization and resensitization of proteinase-activated receptor-2.
abstract: Proteinase-activated receptor-2 (PAR-2) is a G-protein-coupled receptor that is expressed by intestinal epithelial cells, which are episodically exposed to pancreatic trypsin in the intestinal lumen. Trypsin cleaves PAR-2 to expose a tethered ligand, which irreversibly activates the receptor. Thus, PAR-2 may desensitize and resensitize by novel mechanisms. We examined these mechanisms in kidney epithelial cells, stably expressing human PAR-2, and intestinal epithelial cells, which naturally express PAR-2. Trypsin stimulated a prompt increase in [Ca2+]i, due to mobilization of intracellular Ca2+, followed by a sustained plateau, due to influx of extracellular Ca2+. Repeated application of trypsin caused marked desensitization of this response, which is due in part to (a) irreversible cleavage of the receptor by trypsin and (b) protein kinase C-mediated termination of signaling. Trypsin exposure resulted in internalization of PAR-2 into early endosomes and then lysosomes; but endocytosis was not the mechanism of rapid desensitization. Thus, activated PAR-2 is endocytosed and degraded. The Ca2+ response to trypsin resensitized by 60-90 min. Brefeldin A, which disrupted Golgi stores of PAR-2, and cycloheximide, which inhibited protein synthesis, markedly attenuated resensitization. Thus, PAR-2-mediated Ca2+ mobilization desensitizes by irreversible receptor cleavage, protein kinase C-mediated termination of signaling, and PAR-2 targeting to lysosomes. It resensitizes by mobilization of large Golgi stores and synthesis of new receptors.
authors.affiliation: Department of Surgery, University of California, San Francisco, California 94143-0660, USA.
authors.forename: S K
authors.initials: SK
authors.lastname: Böhm
authors.forename: L M
authors.initials: LM
authors.lastname: Khitin
authors.forename: E F
authors.initials: EF
authors.lastname: Grady
authors.forename: G
authors.initials: G
authors.lastname: Aponte
authors.forename: D G
authors.initials: DG
authors.lastname: Payan
authors.forename: N W
authors.initials: NW
authors.lastname: Bunnett
mesh_terms: D001152:Arsenicals / Q000494:pharmacology; D001920:Bradykinin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D002460:Cell Line; D004705:Endocytosis; D005110:Extracellular Space / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D006801:Humans; D016209:Interleukin-8 / Q000494:pharmacology; D007413:Intestinal Mucosa / Q000378:metabolism; D007668:Kidney / Q000378:metabolism; D008247:Lysosomes / Q000737:chemistry; D018613:Microscopy, Confocal; D011493:Protein Kinase C / Q000378:metabolism; D044464:Receptor, PAR-2; D011956:Receptors, Cell Surface / Q000378:metabolism*; D013729:Terpenes / Q000494:pharmacology; D019284:Thapsigargin; D014357:Trypsin / Q000378:metabolism; D010738:Type C Phospholipases / Q000378:metabolism
chemical_list: D001152:Arsenicals; D016209:Interleukin-8; D044464:Receptor, PAR-2; D011956:Receptors, Cell Surface; D013729:Terpenes; C029341:oxophenylarsine; D019284:Thapsigargin; D011493:Protein Kinase C; D010738:Type C Phospholipases; D014357:Trypsin; D000252:Calcium-Transporting ATPases; D001920:Bradykinin; D002118:Calcium