Hiraki Y, Inoue H, Kondo J, Kamizono A, Yoshitake Y, Shukunami C, Suzuki F
Department of Biochemistry and Calcified Tissue Metabolism, Osaka University Faculty of Dentistry, Osaka 565, Japan.
J Biol Chem. 1996 Sep 13;271(37):22657-62. doi: 10.1074/jbc.271.37.22657.
During endochondral bone formation, cartilage cells show increased matrix synthesis and rapid proliferation. We found that cartilage matrix contains at least two types of heparin binding growth-promoting components. One, with a higher affinity to heparin, was identified as chondromodulin I (Hiraki, Y., Tanaka, H., Inoue, H. , Kondo, J., Kamizono, A., and Suzuki, F. (1991) Biochem. Biophys. Res. Commun. 175, 871-977). In this study, we isolated a novel growth-promoting component, chondromodulin II, which has a lower heparin affinity, from the dissociative extracts of fetal bovine epiphyseal cartilage. Chondromodulin II stimulated the proteoglycan synthesis in rabbit cultured growth plate chondrocytes, an expression of the differentiated phenotype of chondrocytes. It also stimulated DNA synthesis in chondrocytes in both the absence and the presence of fibroblast growth factor-2. The apparent molecular mass of chondromodulin II on SDS-polyacrylamide gel electrophoresis was 16 kDa. Its complete amino acid sequence was determined by overlapping sequences of the peptides released by endopeptidase digestion and CNBr cleavage. Chondromodulin II consists of 133 amino acids (calculated Mr = 14,548). The sequence was unique but homologous to the repeats 1 and 2 of the deduced amino acid sequence of the chicken mim-1 gene, which is specifically transactivated by the v-Myb oncogene product in promyelocytes. We also found a minor component with a higher heparin affinity, chondromodulin III, in cartilage extracts. Chondromodulin III stimulated DNA synthesis in chondrocytes in vitro, and its N-terminal sequence was identical with ribosomal protein L31 lacking the N-terminal three amino acids. These findings suggest that the growth and differentiation of chondrocytes are regulated by multiple components in the cartilage matrix.
在软骨内骨形成过程中，软骨细胞表现出基质合成增加和快速增殖。我们发现软骨基质至少含有两种肝素结合生长促进成分。其中一种对肝素亲和力较高的成分被鉴定为软骨调节素I（平木洋、田中浩、井上浩、近藤纯、神园昭、铃木芳，《生物化学与生物物理研究通讯》，1991年，第175卷，第871 - 977页）。在本研究中，我们从胎牛骨骺软骨的解离提取物中分离出一种新的生长促进成分——软骨调节素II，其对肝素的亲和力较低。软骨调节素II刺激了兔培养生长板软骨细胞中蛋白聚糖的合成，这是软骨细胞分化表型的一种表达。在成纤维细胞生长因子 - 2存在和不存在的情况下，它还刺激了软骨细胞中的DNA合成。在SDS - 聚丙烯酰胺凝胶电泳上，软骨调节素II的表观分子量为16 kDa。通过内肽酶消化和CNBr裂解释放的肽段的重叠序列确定了其完整的氨基酸序列。软骨调节素II由133个氨基酸组成（计算分子量 = 14,548）。该序列独特，但与鸡mim - 1基因推导氨基酸序列的重复1和重复2同源，鸡mim - 1基因在早幼粒细胞中被v - Myb癌基因产物特异性反式激活。我们还在软骨提取物中发现了一种对肝素亲和力较高的次要成分——软骨调节素III。软骨调节素III在体外刺激软骨细胞中的DNA合成，其N端序列与缺少N端三个氨基酸的核糖体蛋白L31相同。这些发现表明软骨细胞的生长和分化受软骨基质中的多种成分调节。
