title: Glycosylation broadens the substrate profile of membrane type 1 matrix metalloproteinase.
abstract: Membrane type 1 matrix metalloproteinase (MT1-MMP) is a collagenolytic enzyme that has been implicated in normal development and in pathological processes such as cancer metastasis. The activity of MT1-MMP is regulated extensively at the post-translational level, and the current data support the hypothesis that MT1-MMP activity is modulated by glycosylation. Enzymatic deglycosylation, site-directed mutagenesis, and lectin precipitation assays were used to demonstrate that MT1-MMP contains O-linked complex carbohydrates on the Thr(291), Thr(299), Thr(300), and/or Ser(301) residues in the proline-rich linker region. MT1-MMP glycoforms were detected in human cancer cell lines, suggesting that MT1-MMP activity may be regulated by differential glycosylation in vivo. Although the autolytic processing and interstitial collagenase activity of MT1-MMP were not impaired in glycosylation-deficient mutants, cell surface MT1-MMP-catalyzed activation of pro-matrix metalloproteinase-2 (proMMP-2) required proper glycosylation of MT1-MMP. The inability of carbohydrate-free MT1-MMP to activate proMMP-2 was not a result of defective MT1-MMP zymogen activation, aberrant protein stability, or inability of the mature enzyme to oligomerize. Rather, our data support a mechanism whereby glycosylation affects the recruitment of tissue inhibitor of metalloproteinases-2 (TIMP-2) to the cell surface, resulting in defective formation of the MT1-MMP/TIMP-2/proMMP-2 trimeric activation complex. These data provide evidence for an additional mechanism for post-translational control of MT1-MMP activity and suggest that glycosylation of MT1-MMP may regulate its substrate targeting.
authors.affiliation: Department of Cell & Molecular Biology and Division of Hematology/Oncology, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, USA.
authors.forename: Yi I
authors.initials: YI
authors.lastname: Wu
authors.forename: Hidayatullah G
authors.initials: HG
authors.lastname: Munshi
authors.forename: Ratna
authors.initials: R
authors.lastname: Sen
authors.forename: Scott J
authors.initials: SJ
authors.lastname: Snipas
authors.forename: Guy S
authors.initials: GS
authors.lastname: Salvesen
authors.forename: Rafael
authors.initials: R
authors.lastname: Fridman
authors.forename: M Sharon
authors.initials: MS
authors.lastname: Stack
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002236:Carbohydrate Conformation; D002462:Cell Membrane / Q000378:metabolism; D011232:Chemical Precipitation; D002522:Chlorocebus aethiops; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D006026:Glycoside Hydrolases / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D037102:Lectins; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016297:Mutagenesis, Site-Directed; D017510:Protein Folding; D011498:Protein Precursors / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000378:metabolism; D014407:Tumor Cells, Cultured
chemical_list: D037102:Lectins; D011498:Protein Precursors; D019716:Tissue Inhibitor of Metalloproteinase-2; D006026:Glycoside Hydrolases; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases; D020778:Matrix Metalloproteinase 2