Deposition of tropoelastin into the extracellular matrix requires a competent elastic fiber scaffold but not live cells.
The initial steps of elastic fiber assembly were investigated using an in vitro assembly model in which purified recombinant tropoelastin (rbTE) was added to cultures of live or dead cells. The ability of tropoelastin to associate with preexisting elastic fibers or microfibrils in the extracellular matrix was then assessed by immunofluorescence microscopy using species-specific tropoelastin antibodies. Results show that rbTE can associate with elastic fiber components in the absence of live cells through a process that does not depend on crosslink formation. Time course studies show a transformation of the deposited protein from an initial globular appearance early in culture to a more fibrous structure as the matrix matures. Deposition required the C-terminal region of tropoelastin and correlated with the presence of preexisting elastic fibers or microfibrils. Association of exogenously added tropoelastin to the cellular extracellular matrix was inhibited by the addition of heparan sulfate but not chondroitin sulfate sugars. Together, these results suggest that the matrix elaborated by the cell is sufficient for the initial deposition of tropoelastin in the extracellular space and that elastin assembly may be influenced by the composition of sulfated proteoglycans in the matrix.
Kozel
Beth A
BA
Department of Cell Biology and Physiology, Washington University School of Medicine, Campus Box 8228, 660 South Euclid Ave., St. Louis, MO 63110, USA.
Ciliberto
Christopher H
CH
Mecham
Robert P
RP
D000818:Animals; D002417:Cattle; D002460:Cell Line; D002470:Cell Survival; D002807:Chondroitin / Q000494:pharmacology; D004547:Elastic Tissue / Q000378:metabolism*; D004548:Elasticity; D005109:Extracellular Matrix / Q000737:chemistry / Q000378:metabolism*; D006497:Heparan Sulfate / Q000494:pharmacology; D020894:Microfibrils / Q000378:metabolism; D011485:Protein Binding; D008249:Protein-Lysine 6-Oxidase / Q000037:antagonists & inhibitors; D013997:Time Factors; D014333:Tropoelastin / Q000235:genetics / Q000378:metabolism*
D002807:Chondroitin; D006497:Heparan Sulfate; D008249:Protein-Lysine 6-Oxidase; D014333:Tropoelastin