title: Regulation of ATPase activity of transglutaminase 2 by MT1-MMP: implications for mineralization of MC3T3-E1 osteoblast cultures.
abstract: A pro-mineralization function for transglutaminase 2 (TG2) has been suggested in numerous studies related to bone, cartilage, and vascular calcification. TG2 is an enzyme which can perform protein crosslinking functions, or act as a GTPase/ATPase depending upon different stimuli. We have previously demonstrated that TG2 can act as an ATPase in a Ca(2+)-rich environment and that it can regulate phosphate levels in osteoblast cultures. In this study, we investigate the role MT1-MMP in regulating the ATPase activity of TG2. We report that proteolytic cleavage of TG2 by MT1-MMP in vitro results in nearly a 3-fold increase in the ATPase activity of TG2 with a concomitant reduction in its protein-crosslinking activity. We show that MC3T3-E1 osteoblasts secreted full-length TG2 and major smaller fragments of 66 and 56 kDa, the latter having ATP-binding abilities. MT1-MMP inhibition by a neutralizing antibody suppressed mineralization of osteoblast cultures to 35% of control, and significantly reduced phosphate levels in conditioned medium (CM). Furthermore, MT1-MMP inhibition abolished two of TG2 fragments in the cultures, one of which, the 56-kDa fragment, has ATPase activity. Neutralization of MT1-MMP at early phases of mineralization significantly reduced mineral deposition, but had no effect in later phases implying MT1-MMP and TG2 might contribute to the initiation of mineralization. The cleavage of TG2 by MT1-MMP likely occurs on the cell surface/pericellular matrix where MT1-MMP and TG2 were co-localized. Based on these data, we propose that MT1-MMP modulates the extracellular function TG2 as part of a regulatory mechanism activates the pro-mineralization function of TG2.
authors.affiliation: Division of Biomedical Sciences, Faculty of Dentistry, McGill University, Montreal, Quebec, Canada.
authors.forename: Yukiko
authors.initials: Y
authors.lastname: Nakano
authors.forename: Jennifer
authors.initials: J
authors.lastname: Forsprecher
authors.forename: Mari T
authors.initials: MT
authors.lastname: Kaartinen
mesh_terms: D016475:3T3 Cells; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D002113:Calcification, Physiologic; D002118:Calcium / Q000378:metabolism; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D053511:Matrix Metalloproteinase 14 / Q000378:metabolism*; D051379:Mice; D008970:Molecular Weight; D010006:Osteoblasts / Q000201:enzymology*; D010446:Peptide Fragments; D010710:Phosphates / Q000378:metabolism; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011499:Protein Processing, Post-Translational; D013997:Time Factors; D011503:Transglutaminases / Q000737:chemistry / Q000378:metabolism*
chemical_list: C507096:Mmp14 protein, mouse; D010446:Peptide Fragments; D010710:Phosphates; D000255:Adenosine Triphosphate; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases; D053511:Matrix Metalloproteinase 14; D019204:GTP-Binding Proteins; D002118:Calcium