Regulation of collagen post-translational modification in transformed human and chick-embryo cells

Abstract

Changes in the regulation of collagen post-translational modification in transformed cells were studied in three established human sarcoma cell lines and in chick-embryo fibroblasts freshly transformed by Rous sarcoma virus. The collagens synthesized by all but one of these and by all the control human and chick-embryo cell lines were almost exclusively of types I and/or III. The relative rate of collagen synthesis and the amounts of prolyl hydroxylase activity and immunoreactive protein were markedly low in all the transformed human cell lines. The other enzymes studied, lysyl hydroxylase, hydroxylysyl galactosyltransferase and galactosylhydroxylysyl glucosyltransferase, never showed as large a decrease in activity as did prolyl hydroxylase, suggesting a more efficient regulation of the last enzyme than of the three others. The chick-embryo fibroblasts freshly transformed by Rous sarcoma virus differed from the human sarcoma cells in that prolyl hydroxylase activity was distinctly increased, whereas the decreases in immunoreactive prolyl hydroxylase protein and the three other enzyme activities were very similar to those in the simian-virus-40-transformed human fibroblasts. It seems possible that this increased prolyl hydroxylase activity is only a temporary phenomenon occurring shortly after the transformation, and may be followed by a decrease in activity later. The newly synthesized collagens of all the transformed cells that produced almost exclusively collagen types I and/or III had high extents of lysyl hydroxylation, and there was also an increase in the ratio of glycosylated to non-glycosylated hydroxylysine. The data suggest that one critical factor affecting modification is the rate of collagen synthesis, which affects the ratio of enzyme to substrate in the cell.

Abnormal type I collagen metabolism by cultured fibroblasts in lethal perinatal osteogenesis imperfecta

“…The a I (I)'and a2(I)'-chains in both groups were shown to contain higher levels of hydroxylysine than the normal acl(I)or a2(I)-chains from either control cells or the first group of OI patients (01 26, 30, 31 and 35). The increase in lysine hydroxylation in the al (I)'and a2(I)'-chains was particularly significant because of the already high levels in control al(I)-chains (35%) and a2(I)-chains (52%), as previously reported for cultured fibroblasts (Myllyla et al, 1981). The relative increasein hydroxylation is less marked in the a2(I)'-chain than in the a1(I)'-chain and this may indicate that in cell culture the lysine in the a2(I)-chain is close to maximally hydroxylated.…”

Deficient production of lysyi oxidase in cultures of malignantly transformed human cells

“…Prolyl 4-hydroxylase activity in the 5 sarcoma cell lines studied here is about 25-65% of that in a number of control fibroblast lines [5-71, the activity in the SV40 virus-transformed WI-38 cells being about 40% of that in the non-transformed b Mean + SD WI-38 cells [5]. The activities of the 4 other specific intracellular enzymes of collagen synthesis vary from about 50 to 220% of the control values, part of this variation probably reflecting differences in the types of collagen synthesized .…”

“…The cells were grown in Eagle's minimal essential medium supplemented with 10% foe&l calf serum, 100 units/ml penicillin, 100 @g/ml streptomycin and 50pglml ascorbate [5]. For the measurement of lysyl oxidase activity, subconfluent cells were washed twice with a solution of 0.01 M sodium phosphate and 0.14 M NaCl, pH 7.4, and then cultured for an additional 24 h period in the above medium without serum but supplemented with 5 mg/ml bovine serum albumin [14,17].…”

Collagen fibrillogenesis in tendon development: Current models and regulation of fibril assembly

“…These alterations include the glycosylation of some of the hydroxylysine residues with galactose or glucosylgalactose, and the hydroxylation of about 100 proline residues as well as 5-10 lysine residues to hydroxyproline and hydroxylysine (Torreblanco et al, 1992;Myllyharju, 2005). Many studies have verified a direct relationship between the rate of procollagen folding and the rate of PTMs (Myllyla et al, 1981;Canty and Kadler, 2005). The PTMs begin as soon as the nascent procollagen chains pass into the cisternae of the rough ER and continue until the protein folds into a triple-helical conformation (Torreblanco et al, 1992;Koide and Nagata, 2005).…”