@article {Mukhin1033, author = {Yurii V. Mukhin and Monika Gooz and John R. Raymond and Maria N. Garnovskaya}, title = {Collagenase-2 and -3 Mediate Epidermal Growth Factor Receptor Transactivation by Bradykinin B2 Receptor in Kidney Cells}, volume = {318}, number = {3}, pages = {1033--1043}, year = {2006}, doi = {10.1124/jpet.106.104000}, publisher = {American Society for Pharmacology and Experimental Therapeutics}, 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, Ca2+, calmodulin, protein kinase C, Gαi subunits, or EGFR-B2 receptor heterodimerization. In this study, we demonstrated the involvement of matrix metalloproteinases (MMPs) in B2 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 \~{}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 \~{}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 \~{}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 Gq-coupled BK B2 receptor in mIMCD-3 cells. The American Society for Pharmacology and Experimental Therapeutics}, issn = {0022-3565}, URL = {https://jpet.aspetjournals.org/content/318/3/1033}, eprint = {https://jpet.aspetjournals.org/content/318/3/1033.full.pdf}, journal = {Journal of Pharmacology and Experimental Therapeutics} }