Alignment of tropoelastin molecules during the process of elastogenesis is thought to require fibrillin-containing microfibrils. In this study, we have demonstrated that amino-terminal domains of two microfibrillar proteins, fibrillin-1 and fibrillin-2, interact with tropoelastin in solid phase binding assays. The tropoelastin-binding site was localized to a region beginning at the glycine-rich and proline-rich regions of fibrillin-2 and fibrillin-1, respectively, and continuing through the second 8-cysteine domain. Characterization of the binding requirements using the fibrillin-2 construct found that a folded, secondary structure was necessary for binding. Furthermore, binding between tropoelastin and fibrillin was mediated by ionic interactions involving the lysine side chains of tropoelastin. The importance of the lysine side chains was corroborated by the finding that the fibrillin-2 construct did not bind to mature elastin, whose lysine side chains have been modified to form cross-links. Interestingly, there was no interaction between the fibrillin constructs and tropoelastin in solution phase, suggesting that binding of tropoelastin to a solid substrate exposes a cryptic binding site. These results suggest that fibrillin plays an important role in elastic fiber assembly by binding tropoelastin and perhaps facilitating side chain alignment for efficient cross-linking.

Fibrillin-1 and -2 are large modular extracellular matrix glycoproteins found in many vertebrate organ systems and are known to be key components of the elastic fibre. In the present study, we identify a new heparin-binding region in fibrillin-2 between exons 18 and 24. Additionally, we have narrowed the location of heparin-binding activity previously identified in fibrillin-1 to the last 17 residues of the mature proteolytically processed protein. This domain demonstrated higher activity as a multimer than as a monomer. The fibrillin-1 C-terminal site supported cell attachment in each of nine cell types tested. Attachment was shown to be mediated by cell-surface heparan sulphate proteoglycans. Fibrillin-1 has been shown previously to have heparin-binding activity that is important for matrix deposition of the molecule by fibroblasts. This function in deposition was confirmed in two additional fibrillin-producing cell types (osteosarcoma and epithelial cells) for the deposition of both fibrillin-1 and -2 into the extracellular matrix.

Within tendon, between collagen fascicles, cells are organized in linear arrays surrounded by a specialized environment of extracellular matrix (ECM) proteins that are largely unidentified. Our goal was to identify interfascicular, pericellular ECM components and provide additional resolution to the organization of the pericellular matrix. To this end, we employed a combination of enzymatic digestion, mechanical disruption, and differential sedimentation to demonstrate for the first time that it possible to liberate living linear tendon cell arrays from whole tendon. Here, we identify type VI collagen, versican, and fibrillin-2 as components of the immediate pericellular ECM of linearly arrayed tendon cells. Additionally, a unique fibrillin-2-containing macromolecular assembly is described in detail for the first time. This new structure is unlike any previously described fibrillin-containing macromolecular assembly. Having a largely constant diameter, it runs axially along tendon cell arrays and can exceed 1000 microm in length.

The elastic fiber is known to be an important component of skin, lung, and vasculature. Much less is known about the distribution of elastin and elastic fiber-related proteins in connective tissues, yet genetic defects of elastic fiber constituents can lead to deficiencies in these tissues. For the first time, we determine the distribution of elastin, fibrillins 1 and 2, and microfibril-associated glycoproteins (MAGPs) 1 and 2 in the flexor digitorum profundus (FDP) tendon. Three functionally distinct regions of the FDP tendon, the fibrocartilagenous (FC) region, avascular/tensional (AV/T) region, and insertion region, were evaluated by immunohistochemical methods for these five proteins. Biochemical analysis of desmosine content, an elastin-specific cross-link, demonstrated the presence of elastin in each region, and this was verified histochemically. The fibrillins were found with elastin and also pericellularly with internal fibroblasts where elastin was not detected. Although there was overlapping distribution, fibrillin 2 was more prominent in the interior of the tendon while fibrillin 1 was prominent in outer cell layers that contained elastic fibers. Both MAGP-1 and -2 were found throughout the tendon, although the greatest abundance was near the tendon insertion to bone. Surprisingly, MAGP-1 demonstrated a filamentous appearance within the fibrocartilage that did not correspond to the fibrillin 1 or 2 or MAGP-2 staining pattern. Lastly, we have shown that a vincular membrane located along the dorsal surface of the tendon near the insertion has a very high elastin content and a unique interface with the tendon that consists of an elastic anchor within the tendon body. Anat Rec 268: 430 -440, 2002.