O-Glycosylation Regulates Autolysis of Cellular Membrane Type-1 Matrix Metalloproteinase (MT1-MMP)
Journal of Biological Chemistry
MT1-MMP is a key enzyme in cancer cell invasion and metastasis. The activity of cellular MT1-MMP is regulated by furin-like proprotein convertases, TIMPs, shedding, autoproteolysis, dimerization, exocytosis, endocytosis, and recycling. Our data demonstrate that, in addition to these already known mechanisms, MT1-MMP is regulated by O-glycosylation of its hinge region. Insignificant autolytic degradation is characteristic for naturally expressed, glycosylated, MT1-MMP. In turn, extensive
... , extensive autolytic degradation, which leads to the inactivation of the protease and the generation of its C-terminal membrane-tethered degraded species, is a feature of overexpressed MT1-MMP. We have determined that incomplete glycosylation stimulates extensive autocatalytic degradation and self-inactivation of MT1-MMP. Self-proteolysis commences during the secretory process of MT1-MMP through the cell compartment to the plasma membrane. The strongly negatively charged sialic acid is the most important functional moiety of the glycopart of MT1-MMP. We hypothesize that sialic acid of the O-glycosylation cassette restricts the access of the catalytic domain to the hinge region and to the autolytic cleavage site and protects MT1-MMP from autolysis. Overall, our results point out that there is a delicate balance between glycosylation and self-proteolysis of MT1-MMP in cancer cells and that when this balance is upset the catalytically potent MT1-MMP pool is self-proteolyzed. Membrane-tethered MT1-MMP, 2 the most abundant member of the membrane-type (MT) matrix metalloproteinase subfamily, is distinguished from soluble MMPs by a short transmembrane domain and a cytoplasmic tail (1, 2). MT1-MMP functions in cancer cells as an important mediator of proteolytic events on the cell surface (3, 4), and it is directly engaged in the cleavage of cell surface receptors and the pericellular proteolysis of the extracellular matrix components (5-7). MT1-MMP expression is associated with a variety of pathophysiological conditions and especially with cell locomotion, tumor growth, and metastasis (4, 5, 8 -11). . 2 The abbreviations used are: MT1-MMP, membrane type-1 matrix metalloproteinase; BGN, benzyl 2-acetamido-2-deoxy-␣-D-galactopyranoside; Con A, concanavalin A; dec-RVKR-cmk, decanoyl-Arg-Val-Lys-Arg-chloromethylketone; DMEM, Dulbecco's modified Eagle's medium; FBS, fetal bovine serum; HRP, horseradish peroxidase; MESNA, 2-mercaptoethane sulfonic acid; MMP-2, matrix metalloproteinase-2; PC, proprotein convertase; PDX, ␣ 1 -anti-trypsin variant Portland; TIMP, tissue inhibitor of matrix metalloproteinases; RIPA, radioimmune precipitation assay buffer; PBS, phosphate-buffered saline.