The post-translational phenotype of collagen synthesized by SAOS-2 osteosarcoma cells

Fernandes, Russell J.; Harkey, Michael A.; Weis, MaryAnn; Askew, Jennifer W.; Eyre, David R.

doi:10.1016/j.bone.2007.01.011

Cited by 28 publications

(37 citation statements)

References 56 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…36 The phenotype of the Saos-2 cell is considered to have a mature osteoblast phenotype because it reproduces many of the primary human 37 Saos-2 cells are also able to produce collagenous mineralized bone matrix containing type I and type V collagen and chondroitin sulfate proteoglycans. [38][39][40] Being from a cell line, Saos-2 cells are available for experiments in high numbers, and they are therefore ideal for studies of ECM deposition for which a hyperconfluent layer of cells is needed. In addition, the Saos-2 cell line is advantageous because of its phenotype stability during multiple passages.…”

Section: Discussionmentioning

confidence: 99%

Osteogenic cell differentiation on H-terminated and O-terminated nanocrystalline diamond films

Lišková¹,

Babchenko²,

Varga³

et al. 2015

IJN

View full text Add to dashboard Cite

Nanocrystalline diamond (NCD) films are promising materials for bone implant coatings because of their biocompatibility, chemical resistance, and mechanical hardness. Moreover, NCD wettability can be tailored by grafting specific atoms. The NCD films used in this study were grown on silicon substrates by microwave plasma-enhanced chemical vapor deposition and grafted by hydrogen atoms (H-termination) or oxygen atoms (O-termination). Human osteoblast-like Saos-2 cells were used for biological studies on H-terminated and O-terminated NCD films. The adhesion, growth, and subsequent differentiation of the osteoblasts on NCD films were examined, and the extracellular matrix production and composition were quantified. The osteoblasts that had been cultivated on the O-terminated NCD films exhibited a higher growth rate than those grown on the H-terminated NCD films. The mature collagen fibers were detected in Saos-2 cells on both the H-terminated and O-terminated NCD films; however, the quantity of total collagen in the extracellular matrix was higher on the O-terminated NCD films, as were the amounts of calcium deposition and alkaline phosphatase activity. Nevertheless, the expression of genes for osteogenic markers – type I collagen, alkaline phosphatase, and osteocalcin – was either comparable on the H-terminated and O-terminated films or even lower on the O-terminated films. In conclusion, the higher wettability of the O-terminated NCD films is promising for adhesion and growth of osteoblasts. In addition, the O-terminated surface also seems to support the deposition of extracellular matrix proteins and extracellular matrix mineralization, and this is promising for better osteoconductivity of potential bone implant coatings.

show abstract

Section: Discussionmentioning

confidence: 99%

Osteogenic cell differentiation on H-terminated and O-terminated nanocrystalline diamond films

Lišková¹,

Babchenko²,

Varga³

et al. 2015

IJN

View full text Add to dashboard Cite

show abstract

“…In mature bone, the α1(XI) collagen chain appears and increases with age as a component of type V collagen, presumably in the form of hybrid molecules trimerized with α1(V) and α2(V) chains (Niyibizi and Eyre, 1989). Recent studies on the osteoblast-like cell line, SAOS-2, show expression of α1(XI) and α2(XI) chains in addition to collagen V chains (Fernandes et al, 2007).…”

Section: Discussionmentioning

confidence: 99%

“…After Coomassie Blue staining on SDS-PAGE, individual protein bands were digested in-gel by trypsin. The resulting peptides were subjected to microbore column liquid chromatography (μLC) interfaced directly to a tandem mass spectrometer equipped with a micro-electrospray ionization source (Fernandes et al, 2007). For protein identification, peptide fragments were compared with the NCBI non-redundant protein database using SEQUEST, an automated database search algorithm designed for use with tandem mass spectrometry data.…”

Section: Mass Spectrometrymentioning

confidence: 99%

Collagen XI chain misassembly in cartilage of the chondrodysplasia (cho) mouse

et al. 2007

Self Cite

View full text Add to dashboard Cite

Molecular mechanisms controlling the assembly of cartilage-specific types II, IX and XI collagens into a heteropolymeric network of uniformly thin, unbanded fibrils are not well understood, but collagen XI has been implicated. The present study on cartilage from the homozygous chondrodysplasia (cho/cho) mouse adds support to this concept. In the absence of α1(XI) collagen chains, thick, banded collagen fibrils are formed in the extracellular matrix of cho/cho cartilage. A functional knock-out of the type XI collagen molecule has been assumed. We have re-examined this at the protein level to see if rather than a complete knock-out, alternative type XI chain assemblies were formed. Mass spectrometry of purified triple helical collagen from the rib cartilage of cho/ cho mice identified α1(V) and α2(XI) chains. These chains were recovered in roughly equal amounts based on Coomassie blue staining of SDS-PAGE gels, in addition to α1(II)/α3(XI) collagen chains. Using telopeptide-specific antibodies and Western blot analysis, it was further shown that type V/ XI trimers were present in the matrix cross-linked to each other and to type II collagen molecules to form heteropolymers. Cartilage from heterozygous (cho/+) mice contained a mix of α1(V) and α1 (XI) chains and a mix of thin and thick fibrils on transmission electron microscopy. In summary, the results imply that native type XI collagen molecules containing an α1(XI) chain are required to form uniformly thin fibrils and support a role for type XI collagen as the template for the characteristic type II collagen fibril network of developing cartilage.

show abstract

“…The SAOS-2 cell line (ATCC catalog no. HTB-85) was maintained as monolayer cultures for a month in McCoy's medium containing 10% FBS (Invitrogen) and 50 g/ml ascorbate (23,24). The human chondrosarcoma cell line, CH1.2 (24), human breast cancer cell lines MDA 231 (ATCC catalog no.…”

Section: Methodsmentioning

confidence: 99%

A Role for Prolyl 3-Hydroxylase 2 in Post-translational Modification of Fibril-forming Collagens

Fernandes

Farnand

Traeger

et al. 2011

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

The fibrillar collagen types I, II, and V/XI have recently been shown to have partially 3-hydroxylated proline (3Hyp) residues at sites other than the established primary Pro-986 site in the collagen triple helical domain. These sites showed tissue specificity in degree of hydroxylation and a pattern of D-periodic spacing. This suggested a contributory role in fibril supramolecular assembly. The sites in clade A fibrillar ␣1(II), ␣2(V), and ␣1(I) collagen chains share common features with known prolyl 3-hydroxylase 2 (P3H2) substrate sites in ␣1(IV) chains implying a role for this enzyme. We pursued this possibility using the Swarm rat chondrosarcoma cell line (RCS-LTC) found to express high levels of P3H2 mRNA. Mass spectrometry determined that all the additional candidate 3Hyp substrate sites in the pN type II collagen made by these cells were highly hydroxylated. In RNA interference experiments, P3H2 protein synthesis was suppressed coordinately with prolyl 3-hydroxylation at Pro-944, Pro-707, and the C-terminal GPP repeat of the pN␣1(II) chain, but Pro-986 remained fully hydroxylated. Furthermore, when P3H2 expression was turned off, as seen naturally in cultured SAOS-2 osteosarcoma cells, full 3Hyp occupancy at Pro-986 in ␣1(I) chains was unaffected, whereas 3-hydroxylation of residue Pro-944 in the ␣2(V) chain was largely lost, and 3-hydroxylation of Pro-707 in ␣2(V) and ␣2(I) chains were sharply reduced. The results imply that P3H2 has preferred substrate sequences among the classes of 3Hyp sites in clade A collagen chains.Collagen, the major structural protein of vertebrates has evolved a range of post-translational modifications that are essential for triple helix assembly and stability, intermolecular cross-linking, and strength of fibrils and tissue function. Enzymes from the family of 2-oxo-glutarate-dependent dioxygenases are responsible for most of these modifications. For example, prolyl 4-hydroxylase catalyzes proline 4-hydroxylation, a modification necessary for the secondary and tertiary structure of collagen (1, 2), and the lysyl hydroxylase isoenzymes catalyze lysine hydroxylation, a modification essential for the formation of intermolecular cross-links in collagen (2-4).Prolyl 3-hydroxlase 1 (P3H1), 2 another member of the 2-oxo-glutarate-dependent dioxygenase family, catalyzes the post-translational 3-hydroxylation of certain proline residues in fibril-forming collagens. This enzyme was only recently cloned from chicken tissues (5) after being partially purified Ͼ25 years ago (6). P3H1 was identified as the chick homologue of rat leprecan, a glycoprotein, which was first isolated from a rat parietal yolk sac tumor cell line (7). Within the past few years, novel mutations in the human P3H1 gene and genes encoding P3H1-associated proteins were shown to cause recessive forms of osteogenesis imperfecta (8 -11). By mass spectrometry, we and other investigators have shown that Pro-986 in the ␣1(I) collagen chain of bone and skin from such patients can be severely under-3-hydroxyated. The P3H1 enz...

show abstract

The post-translational phenotype of collagen synthesized by SAOS-2 osteosarcoma cells

Cited by 28 publications

References 56 publications

Osteogenic cell differentiation on H-terminated and O-terminated nanocrystalline diamond films

Osteogenic cell differentiation on H-terminated and O-terminated nanocrystalline diamond films

Collagen XI chain misassembly in cartilage of the chondrodysplasia (cho) mouse

A Role for Prolyl 3-Hydroxylase 2 in Post-translational Modification of Fibril-forming Collagens

Contact Info

Product

Resources

About