EFFECT OF RETINOIC ACID ON PROTEIN-SYNTHESIS BY FETAL BOVINE CHONDROCYTES IN HIGH-DENSITY CULTURE - DOWN-REGULATION OF THE GLUCOSE-REGULATED PROTEIN, GRP-78, AND TYPE-II COLLAGEN
Am. Freyria et al., EFFECT OF RETINOIC ACID ON PROTEIN-SYNTHESIS BY FETAL BOVINE CHONDROCYTES IN HIGH-DENSITY CULTURE - DOWN-REGULATION OF THE GLUCOSE-REGULATED PROTEIN, GRP-78, AND TYPE-II COLLAGEN, Biochemical journal, 305, 1995, pp. 391-396
The effect of 0.1-10 mu M retinoic acid (RA) on foetal bovine chondroc
ytes was investigated in high-density cultures (0.6 x 10(6) cells/cm(2
)). After 5 days of culture in ascorbate-free medium, control chondroc
ytes presented a typical rounded shape and synthesized type II, IX, XI
and III collagens. After RA treatment on days 2-5 of culture, the cel
ls exhibited a fibroblast-like shape and decreased synthesis of total
protein (48%) and pepsin-resistant proteins (60%) as determined by [S-
35]methionine labelling. Addition of RA was not followed by the expres
sion of type I collagen, but induced quantitative changes in the synth
esis of cartilage-specific collagens (II, IX and XI) as measured by di
rect autoradiography of the corresponding bands after SDS/PAGE. The ma
in change was in type II collagen synthesis, with a 80% decrease in th
e cell-layer fraction and a 89% decrease in culture-medium fraction; i
nhibition of type IX and XI collagen synthesis was limited to 25 and 3
1% respectively. Modifications to intracellular proteins induced by RA
were determined by using two-dimensional electrophoresis associated w
ith a computerized imaging system. Synthesis of one of the more abunda
nt proteins (pI 4.8; 78 kDa) was decreased by 75% after RA treatment.
This protein was characterized by micro-sequencing as the glucose-regu
lated protein 78 (GRP 78). It was reported previously to bind denature
d collagen and mutated type I procollagen molecule and to function as
a molecular chaperone for collagen molecules. It remains to demonstrat
e whether the parallel down-regulation of GRP 78 and type II collagen
observed here corresponds to a co-ordinate regulation of these two pro
teins.