Molecular assembly, secretion, and matrix deposition of type VI collagen.

Abstract

Abstract. Monoclonal antibodies reactive with the tissue form of type VI collagen were used to isolate the type VI collagen polypeptides from cultured fibroblasts and muscle cells. Two [35S]methionine-labeled polypeptides of 260 and 140 kD were found intracellularly, in the medium, and in the extracellular matrix of metabolically labeled cells. These polypeptides were disulfide cross-linked into very large complexes. The 260-and 140-kD polypeptides were intimately associated and could not be separated from each other by reduction without denaturation. In the absence of ascorbic acid, both polypeptides accumulated inside the cell, and their amounts in the medium and in the matrix were decreased. These results suggest that both the 260-and the 140-kD polypeptides are integral parts of the type VI collagen molecule. Examination of type VI collagen isolated from the intracellular pool by electron microscopy after rotary shadowing revealed structures corresponding to different stages of assembly of type VI collagen. Based on these images, a sequence for the intracellular assembly of type VI collagen could be discerned. Type VI collagen monomers are ~ 125 nm long and are composed of two globules separated by a thin strand. The monomers assemble into dimers and tetramers by lateral association. Only tetramers were present in culture media, whereas both tetramers and multimers were found in extracellular matrix extracts. The multimers appeared to have assembled from tetramers by end-to-end association into filaments that had prominent knobs and a periodicity of-110 nm. These results show that, unlike other collagens, type VI collagen is assembled into tetramers before it is secreted from the cells, and they also suggest an extracellular aggregation mechanism that appears to be unique to this collagen.T YPE VI collagen is widely distributed throughout connective tissues (l 3, 24). It has been isolated from aorta and placenta in the form of high molecular weight, pepsin-resistant fragments composed of 50-80-kD polypeptides (3,8,9,19,13,14,16,19,21). It has also been isolated from these tissues by guanidine hydrochloride extraction as a protein composed of polypeptides ranging from 140 kD (10, 15) up to 190 kD (21). One outstanding feature of both the pepsin fragments and the larger forms of type VI collagen is the extensive interchain disulfide cross-linking. It appears from chemical and morphological analyses of type VI collagen that it contains relatively short triple-helical domains, contributing one-third or less to the total mass of the protein (7,10,13,15,19).The molecular architecture (2, 7, 13, 15), the large amount of globular structure, and the extensive disulfide cross-linking in type VI collagen suggest that it is unique among collagens, but little is known about its biosynthesis, processing, and biological function. Type VI collagen is synthesized by fibroblasts and muscle cells in culture and is deposited into their extracellular matrix (l l, 13, 22, 24). The type VI collagen immunoreactivity in cell cultur...

Extracellular matrix in the trabecular meshwork

“…4. (Bruns et al 1986;Engvall et al 1986;Ball et al 2001;Reale et al 2001;Baldock et al 2003;Ball et al 2003;Knupp et al 2006) In the TM, type VI immunostaining of both heteromorphic aggregates and interband longitudinal sheets has been observed, supporting the model in Fig. 4.…”

“…(Reale et al 2001) This may relate to the 100 nm repeat structures in the JCT, although this has not been comprehensively studied in detail. (Hessle and Engvall 1984;Bruns et al 1986;Engvall et al 1986;Knupp et al 2006) The type VI collagen N-terminal repeats bind hyaluronan and heparin, while the C-terminal domains are involved in self-assembly. (Kielty et al 1992b;Specks et al 1992;Ball et al 2001;Ball et al 2003) Decorin and biglycan associate with the N-terminal end of the central collagenous domain, affecting fibril formation.…”

Collagens in human atherosclerosis. Immunohistochemical analysis using collagen type-specific antibodies.

“…32 Type VI collagen is a "short-chain" molecule consisting of a triple helical segment with globular domains at each end. 40 - 42 The end-on-end aggregation of tetramers gives rise to filaments that have been observed in cell culture as beaded filaments, with the beads representing a structure formed by the interaction of globular domains from two tetramers. 40 ' 43 The beaded filaments described in our study appear to be similar to those described by Brans et al 35 and Keene et al 33 However, Keene et al 33 have shown that type VI collagen is a component of a ruthenium red-stainable network.…”

Collagen type VI in neural crest development: Distribution in situ and interaction with cells in vitro

“…The triple-helical collagenous portion of the molecule is relatively short and is flanked by extended noncollagenous domains, which after rotary shadowing appear as prominent globules. These terminal globules are thought to be important for the macromolecular assembly of the molecule (Furthmayr et al, 1983;von der Mark et al, 1984;Jander et al, 1984;Engvall et al, 1986;Kuo et al, 1989). Monomers of Col VI associate intracellularly to form disulfidestabilized tetramers which organize into linear polymeric complexes after secretion from the cell.…”