Changes in heparan sulfate are associated with delayed wound repair, altered cell migration, adhesion and contractility in the galactosyltransferase I (ß4GalT-7) deficient form of Ehlers–Danlos syndrome

• Published in: Human molecular genetics 2008-04, Vol.17 (7), p.996-1009
• Main Authors: Götte, Martin, Spillmann, Dorothe, Yip, George W., Versteeg, Elly, Echtermeyer, Frank G., van Kuppevelt, Toin H., Kiesel, Ludwig
• Format: Article
• Language: English
• Subjects: Actins - metabolism; Animals; Biological and medical sciences; Caenorhabditis elegans; Cell Adhesion; Cell Movement; Drosophila melanogaster; Ehlers-Danlos Syndrome - enzymology; Ehlers-Danlos Syndrome - genetics; Ehlers-Danlos Syndrome - physiopathology; Female; Fibrin - physiology; Fibroblasts - cytology; Fibronectins - physiology; Fundamental and applied biological sciences. Psychology; Galactosyltransferases - genetics; Galactosyltransferases - metabolism; Gene Expression Profiling; Genetics of eukaryotes. Biological and molecular evolution; Glycosaminoglycans - biosynthesis; Glycosaminoglycans - genetics; Heparitin Sulfate - chemistry; Heparitin Sulfate - metabolism; Humans; Infant; Male; Medical sciences; MEDICIN; MEDICINE; Mice; Molecular and cellular biology; Pseudopodia - physiology; Reverse Transcriptase Polymerase Chain Reaction; RNA, Small Interfering; Sarcoidosis. Granulomatous diseases of unproved etiology. Connective tissue diseases. Elastic tissue diseases. Vasculitis; Stress Fibers - metabolism; Syndecan-4 - genetics; Syndecan-4 - metabolism; Wound Healing
• ISSN: 0964-6906; 1460-2083; 1460-2083
• DOI: 10.1093/hmg/ddm372

Reduced activity of ß4-galactosyltransferase 7 (ß4GalT-7), an enzyme involved in synthesizing the glycosaminoglycan linkage region of proteoglycans, is associated with the progeroid form of Ehlers–Danlos syndrome (EDS). In the invertebrates Drosophila melanogaster and Caenorhabditis elegans, mutations in ß4GalT-7 affect biosynthesis of heparan sulfate (HS), a modulator of several biological processes relevant to wound repair. We have analyzed structural alterations of HS and their functional consequences in human ß4GalT-7 Arg270Cys mutant EDS and control fibroblasts. HS disaccharide analysis by reversed phase ion-pairing chromatography revealed a reduced sulfation degree of HS paralleled by altered immunostaining patterns for the phage-display anti-HS antibodies HS4E4 and RB4EA12 in ß4GalT-7 mutant fibroblasts. Real-time PCR-analysis of 44 genes involved in glycosaminoglycan biosynthesis indicated that the structural alterations in HS were not caused by differential regulation at the transcriptional level. Scratch wound closure was delayed in ß4GalT-7-deficient cells, which could be mimicked by enzymatic removal of HS in control cells. siRNA-mediated knockdown of ß4GalT-7 expression induced morphological changes in control fibroblasts which suggested altered cell–matrix interactions. Adhesion of ß4GalT-7 deficient cells to fibronectin was increased while actin stress fiber formation was impaired relative to control cells. Also collagen gel contraction was delayed in the ß4GalT-7 mutants which showed a reduced formation of pseudopodia and filopodia, less efficient penetration of the collagen gels and a diminished formation of collagen suprastructures. Our study suggests an HS-dependent basic mechanism behind the altered wound repair phenotype of ß4GalT-7-deficient EDS patients.