title: Collagenase-2 and -3 mediate epidermal growth factor receptor transactivation by bradykinin B2 receptor in kidney cells.
abstract: We have previously shown that stimulation of extracellular signal-regulated protein kinase (ERK) by bradykinin (BK) in murine inner medullary collecting duct (mIMCD)-3 cells is mediated by epidermal growth factor receptor (EGFR) transactivation. The mechanism of EGFR transactivation seemed to be novel, because it does not require phospholipase C, Ca(2+), calmodulin, protein kinase C, G alpha(i) subunits, or EGFR-B(2) receptor heterodimerization. In this study, we demonstrated the involvement of matrix metalloproteinases (MMPs) in B(2) receptor-induced EGFR transactivation using their broad-spectrum inhibitors batimastat and N-[(2R)-2-(hydroxamidocarbonylmethyl)-4-methylpentanoyl]-l-tryptophan methylamide (Galardin) (GM-6001). Selective inhibitors for collagenase-2 and -3 (MMP-8 and MMP-13, respectively) blocked BK-induced EGFR phosphorylation and ERK activation, whereas inhibitors for MMP-1, -2, -3, -7, or -9 were without effect. Transfection of mIMCD-3 cells with MMP-8 small interfering RNA (siRNA) resulted in approximately 50% decrease of BK-induced ERK activation. A neutralizing antibody against MMP-13 as well as transfection with MMP-13 siRNA produced a similar effect. Inhibition of both collagenases resulted in approximately 65% decrease of BK-induced ERK activation, supporting roles for both enzymes. Stimulation of mIMCD-3 cells with 10 nM BK increased the activity of collagenases in concentrated culture media within 10 min. Moreover, recombinant MMP-13 and MMP-8, when applied to mIMCD-3 cells for 10 min without BK, stimulated tyrosine phosphorylation of EGFR and caused approximately 250% increase over basal ERK phosphorylation comparable with BK-induced ERK activation. Collagenases-induced ERK activation was inhibited by 4-(3-chloroanilino)-6,7-dimethoxyquinazoline (AG-1478) and thus dependent on EGFR tyrosine kinase activity. This study demonstrates a novel role for collagenase-2 and -3 in signaling of the G(q)-coupled BK B(2) receptor in mIMCD-3 cells.
authors.affiliation: Ralph H. Johnson Veterans Affairs Medical Center, Department of Medicine, Medical University of South Carolina, Charleston, SC 29425-2227, USA.
authors.forename: Yurii V
authors.initials: YV
authors.lastname: Mukhin
authors.forename: Monika
authors.initials: M
authors.lastname: Gooz
authors.forename: John R
authors.initials: JR
authors.lastname: Raymond
authors.forename: Maria N
authors.initials: MN
authors.lastname: Garnovskaya
mesh_terms: D000818:Animals; D001920:Bradykinin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D017364:Collagenases / Q000502:physiology*; D004789:Enzyme Activation; D066246:ErbB Receptors / Q000378:metabolism*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000378:metabolism*; D053509:Matrix Metalloproteinase 13; D020784:Matrix Metalloproteinase 8 / Q000502:physiology*; D051379:Mice; D010766:Phosphorylation; D034741:RNA, Small Interfering / Q000494:pharmacology; D043782:Receptor, Bradykinin B2 / Q000502:physiology*; D015533:Transcriptional Activation; D016211:Transforming Growth Factor alpha / Q000502:physiology
chemical_list: D034741:RNA, Small Interfering; D043782:Receptor, Bradykinin B2; D016211:Transforming Growth Factor alpha; D066246:ErbB Receptors; D048049:Extracellular Signal-Regulated MAP Kinases; D017364:Collagenases; D053509:Matrix Metalloproteinase 13; C507081:Mmp13 protein, mouse; D020784:Matrix Metalloproteinase 8; D001920:Bradykinin; D002118:Calcium