Heparan sulfate is necessary for the early formation of nascent fibronectin and collagen I fibrils at matrix assembly sites
| Autor: | Katherine E. Hill, Benjamin M. Lovett, Jean E. Schwarzbauer |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
HSPG
heparan sulfate-modified proteoglycan BCS bovine calf serum GAPDH glyceraldehyde 3-phosphate dehydrogenase HS heparan sulfate CHO Cells heparin Biochemistry glycosyltransferase CHO Chinese hamster ovary Collagen Type I Cricetulus fibronectin GAG glycosaminoglycans Cricetinae Animals matrix assembly DOC deoxycholate detergent DTDST diastrophic dysplasia sulfate transporter Molecular Biology FN fibronectin HME hereditary multiple exostoses Cell Biology ECM extracellular matrix Extracellular Matrix Fibronectins heparan sulfate Heparitin Sulfate EXT1 exostosin-1 DMEM Dulbecco’s Modified Eagle’s Medium EXT2 exostosin-2 Research Article |
| Zdroj: | The Journal of Biological Chemistry |
| ISSN: | 1083-351X 0021-9258 |
| Popis: | Fibronectin (FN), an essential component of the extracellular matrix (ECM), is assembled via a cell-mediated process in which integrin receptors bind secreted FN and mediate its polymerization into fibrils that extend between cells, ultimately forming an insoluble matrix. Our previous work using mutant Chinese hamster ovary (CHO) cells identified the glycosaminoglycan heparan sulfate (HS) and its binding to FN as essential for the formation of insoluble FN fibrils. In this study, we investigated the contributions of HS at an early stage of the assembly process using knockdown of exostosin-1 (EXT1), one of the glycosyltransferases required for HS chain synthesis. NIH 3T3 fibroblasts with decreased EXT1 expression exhibited a significant reduction in both FN and type I collagen in the insoluble matrix. We show that FN fibril formation is initiated at matrix assembly sites, and while these sites were formed by cells with EXT1 knockdown, their growth was stunted compared with wild-type cells. The most severe defect observed was in the polymerization of nascent FN fibrils, which was reduced 2.5-fold upon EXT1 knockdown. This defect was rescued by the addition of exogenous soluble heparin chains long enough to simultaneously bind multiple FN molecules. The activity of soluble heparin in this process indicates that nascent fibril formation depends on HS more so than on the protein component of a specific HS proteoglycan. Together, our results suggest that heparin or HS is necessary for concentrating and localizing FN molecules at sites of early fibril assembly. |
| Databáze: | OpenAIRE |
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