Fibrillin proteins constitute the backbone of extra-cellular macromolecular microfibrils. Mutations in
fibrillins cause heritable connective tissue disorders, including
Marfan syndrome, dominant
Weill-Marchesani syndrome, and
stiff skin syndrome.
Fibronectin provides a critical scaffold for microfibril assembly in cell culture models. Full length recombinant
fibrillin-1 was expressed by HEK 293 cells, which deposited the secreted
protein in a punctate pattern on the cell surface. Cocultured fibroblasts consistently triggered assembly of recombinant
fibrillin-1, which was dependent on a
fibronectin network formed by the fibroblasts. Deposition of recombinant
fibrillin-1 on
fibronectin fibers occurred first in discrete packages that subsequently extended along
fibronectin fibers. Mutant
fibrillin-1 harboring either a
cysteine 204 to
serine mutation or a RGD to RGA mutation which prevents
integrin binding, did not affect
fibrillin-1 assembly. In conclusion, we developed a modifiable recombinant full-length
fibrillin-1 assembly system that allows for rapid analysis of critical roles in
fibrillin assembly and functionality. This system can be used to study the contributions of specific residues, domains, or regions of
fibrillin-1 to the biogenesis and functionality of microfibrils. It provides also a method to evaluate disease-causing mutations, and to produce microfibril-containing matrices for tissue engineering applications, for example, in designing novel vascular grafts or
stents.