Proteolysis of cell-surface tissue transglutaminase by matrix metalloproteinase-2 contributes to the adhesive defect and matrix abnormalities in thrombospondin-2-null fibroblasts and mice.
Thrombospondin (TSP)-2-null dermal fibroblasts display an attachment defect that results from increased matrix metalloproteinase (MMP)-2 levels in their conditioned media. To investigate the molecular mechanisms responsible for this defect, we analyzed the activity of tissue transglutaminase (tTG) in TSP-2-null dermal fibroblasts and in tissues of TSP-2-null mice. tTG functions as a co-receptor for beta1 and beta3 integrins and stabilizes extracellular matrix proteins by introduction of isopeptide cross-links. Cell-surface tTG activity was reduced in TSP-2-null cells (0.50 +/- 0.05 arbitrary units versus 0.84 +/- 0.07 for wild type; P < or = 0.05), and addition of MMP-2 to the culture medium of wild-type cells caused a 35% reduction in cell-surface tTG activity. tTG was susceptible to proteolysis by MMP-2 in vitro, and addition of the MMP inhibitor TIMP-2 to TSP-2-null cells restored tTG activity (0.3 +/- 0.08 for untreated cells; 0.71 +/- 0.09 with TIMP-2). TSP-2-null mice had reduced tTG activity in skin, as measured by incorporation of fluorescein isothiocyanate-labeled cadaverine, and a threefold increase in acetic acid-extracted dermal collagen. Furthermore, isopeptide cross-links were reduced in both uninjured skin and in excisional wounds of TSP-2-null mice, as determined by morphometric immunohistochemical analysis, indicating that isopeptide cross-links are important for the stabilization of the collagenous matrix in dermis. These findings provide a mechanism for the reduced adhesion of TSP-2-null fibroblasts and an explanation for the increased collagen solubility and fragility of TSP-2-null skin.
Agah
Azin
A
Department of Biochemistry, School of Medicine, University of Washington, Seattle, Washington, USA.
Kyriakides
Themis R
TR
Bornstein
Paul
P
D000818:Animals; D002448:Cell Adhesion / Q000502:physiology*; D003599:Cytoskeleton / Q000378:metabolism*; D005347:Fibroblasts / Q000201:enzymology*; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D012867:Skin / Q000201:enzymology; D019699:Thrombospondins / Q000235:genetics; D019714:Tissue Inhibitor of Metalloproteinases / Q000494:pharmacology; D011503:Transglutaminases / Q000187:drug effects / Q000378:metabolism*; D000099262:Thrombospondin 2
D020778:Matrix Metalloproteinase 2; D008562:Membrane Glycoproteins; D019699:Thrombospondins; D019714:Tissue Inhibitor of Metalloproteinases; D011503:Transglutaminases; D000099262:Thrombospondin 2