Multiple bone morphogenetic protein 1-related mammalian metalloproteinases process pro-lysyl oxidase at the correct physiological site and control lysyl oxidase activation in mouse embryo fibroblast cultures.
Lysyl oxidase catalyzes the final enzymatic step required for collagen and elastin cross-linking in extracellular matrix biosynthesis. Pro-lysyl oxidase is processed by procollagen C-proteinase activity, which also removes the C-propeptides of procollagens I-III. The Bmp1 gene encodes two procollagen C-proteinases: bone morphogenetic protein 1 (BMP-1) and mammalian Tolloid (mTLD). Mammalian Tolloid-like (mTLL)-1 and -2 are two genetically distinct BMP-1-related proteinases, and mTLL-1 has been shown to have procollagen C-proteinase activity. The present study is the first to directly compare pro-lysyl oxidase processing by these four related proteinases. In vitro assays with purified recombinant enzymes show that all four proteinases productively cleave pro-lysyl oxidase at the correct physiological site but that BMP-1 is 3-, 15-, and 20-fold more efficient than mTLL-1, mTLL-2, and mTLD, respectively. To more directly assess the roles of BMP-1 and mTLL-1 in lysyl oxidase activation by connective tissue cells, fibroblasts cultured from Bmp1-null, Tll1-null, and Bmp1/Tll1 double null mouse embryos, thus lacking BMP-1/mTLD, mTLL-1, or all three enzymes, respectively, were assayed for lysyl oxidase enzyme activity and for accumulation of pro-lysyl oxidase and mature approximately 30-kDa lysyl oxidase. Wild type cells or cells singly null for Bmp1 or Tll1 all produced both pro-lysyl oxidase and processed lysyl oxidase at similar levels, indicating apparently normal levels of processing, consistent with enzyme activity data. In contrast, double null Bmp1/Tll1 cells produced predominantly unprocessed 50-kDa pro-lysyl oxidase and had lysyl oxidase enzyme activity diminished by 70% compared with wild type, Bmp1-null, and Tll1-null cells. Thus, the combination of BMP-1/mTLD and mTLL-1 is shown to be responsible for the majority of processing leading to activation of lysyl oxidase by murine embryonic fibroblasts, whereas in vitro studies identify pro-lysyl oxidase as the first known substrate for mTLL-2.
Uzel
M I
MI
Division of Oral Biology, Boston University Goldman School of Dental Medicine, Boston, Massachusetts 02118, USA.
Scott
I C
IC
Babakhanlou-Chase
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H
Palamakumbura
A H
AH
Pappano
W N
WN
Hong
H H
HH
Greenspan
D S
DS
Trackman
P C
PC
D000818:Animals; D055395:Bone Morphogenetic Protein 1; D019485:Bone Morphogenetic Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D004622:Embryo, Mammalian / Q000166:cytology / Q000201:enzymology*; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology / Q000201:enzymology; D006801:Humans; D008666:Metalloendopeptidases / Q000378:metabolism*; D051379:Mice; D011499:Protein Processing, Post-Translational; D008249:Protein-Lysine 6-Oxidase / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism
D019485:Bone Morphogenetic Proteins; D004792:Enzyme Precursors; D011994:Recombinant Proteins; D008249:Protein-Lysine 6-Oxidase; D008666:Metalloendopeptidases; C525705:BMP1 protein, human; C525707:Bmp1 protein, mouse; D055395:Bone Morphogenetic Protein 1