Regulation Of Fibril Diameter Research Articles

Collagen a1(XI) structure prediction by Alphafold 2.

Collagen α1(XI) is a minor fibrillar collagen involved in the critical regulation of collagen fibrils such as nucleation, assembly, and regulation of fibril diameter. The amino propeptide domain of the collagen α1(XI) is retained on the surface of the collagen fibril for an extended period of time and may play a crucial role in the interaction with extracellular matrix glycosaminoglycans and other proteins during the process of fibrillogenesis. Understanding the mechanism of action of this protein will ultimately help us understand the organization and assembly of the extracellular matrix that underlies the structural integrity of connective tissues.

Type V Collagen Controls the Initiation of Collagen Fibril Assembly

Vertebrate collagen fibrils are heterotypically composed of a quantitatively major and minor fibril collagen. In non-cartilaginous tissues, type I collagen accounts for the majority of the collagen mass, and collagen type V, the functions of which are poorly understood, is a minor component. Type V collagen has been implicated in the regulation of fibril diameter, and we reported recently preliminary evidence that type V collagen is required for collagen fibril nucleation (Wenstrup, R. J., Florer, J. B., Cole, W. G., Willing, M. C., and Birk, D. E. (2004) J. Cell. Biochem. 92, 113-124). The purpose of this study was to define the roles of type V collagen in the regulation of collagen fibrillogenesis and matrix assembly. Mouse embryos completely deficient in pro-alpha1(V) chains were created by homologous recombination. The col5a1-/- animals die in early embryogenesis, at approximately embryonic day 10. The type V collagen-deficient mice demonstrate a virtual lack of collagen fibril formation. In contrast, the col5a1+/- animals are viable. The reduced type V collagen content is associated with a 50% reduction in fibril number and dermal collagen content. In addition, relatively normal, cylindrical fibrils are assembled with a second population of large, structurally abnormal collagen fibrils. The structural properties of the abnormal matrix are decreased relative to the wild type control animals. These data indicate a central role for the evolutionary, ancient type V collagen in the regulation of fibrillogenesis. The complete dependence of fibril formation on type V collagen is indicative of the critical role of the latter in early fibril initiation. In addition, this fibril collagen is important in the determination of fibril structure and matrix organization.

Production in mammalian cells of chimeric human/sea urchin procollagen molecules displaying distinct versions of the minor triple helix.

One of the mechanisms involved in the regulation of the fibril diameter is the retention of the N-propeptide. In sea urchin embryo, thin collagen fibrils harbor numerous extensions at their surface, which we have suggested correspond to the large N-propeptide of the 2alpha collagen chain. To investigate the function of the N-propeptide during fibrillogenesis, we engineered constructs coding for the globular region of the 2alpha N-propeptide. To obtain homotrimeric molecules, the N-telopeptide, the central triple helix and the C-propeptide of the 2alpha chain were replaced by human domains of the proalpha1(I) chain. A single restriction site allowed insertion of distinct versions of the minor triple helix of the N-propeptide. Several human cell lines were transfected, and with one of them we were able to produce intact homotrimeric procollagen molecules. Rotary shadowing of these purified molecules indicates the presence of three large 2alpha N-propeptides that are similar to the extensions present at the surface of the sea urchin thin fibrils. This cassette-vector will be useful in determining the respective contributions of the globular and minor triple helical domains of the N-propeptide in the regulation of fibril diameter.

Collagen fibril assembly by corneal fibroblasts in three-dimensional collagen gel cultures: Small-diameter heterotypic fibrils are deposited in the absence of keratan sulfate proteoglycan

Extracellular matrix assembly is a multistep process and the various steps in collagen fibrillogenesis are thought to be influenced by a number of factors, including other noncollagenous matrix molecules. The synthesis and deposition of extracellular matrix by corneal fibroblasts grown within three-dimensional collagen gel cultures were examined to elucidate the factors important in the establishment of tissue-specific matrix architecture. Corneal fibroblasts in collagen gel cultures form layers and deposit small-diameter collagen fibrils (~25 nm) typical of the mature corneal stroma. The matrix synthesized contains type VI collagen in a filamentous network and type I and type V collagen assembled as heterotypic fibrils. The amount of type V collagen synthesized is relatively high and comparable to that seen in the corneal stroma. This matrix is deposited between cell layers in a manner reminiscent of the secondary corneal stroma, but is not deposited as densely or as organized as would be found in situ. No keratan sulfate proteoglycan, a proteoglycan found only in the corneal stroma, was synthesized by the fibroblasts in the collagen gel cultures. The assembly and deposition of small-diameter fibrils with a collagen composition and structure identical to that seen in the corneal stroma in the absence of proteoglycans typical of the secondary corneal stroma imply that although proteoglycan-collagen interactions may function in the establishment of interfibrillar spacing and lamellar organization, collagen-collagen interactions are the major parameter in the regulation of fibril diameter.