Chondroitin sulfate N-acetylgalactosaminyltransferase-1 plays a critical role in chondroitin sulfate synthesis in cartilage
Chondroitin sulfate N-acetylgalactosaminyltransferase-1 plays a critical role in chondroitin sulfate synthesis in cartilage
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DOI：
10.1074/jbc.m606870200
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发表时间：
2007-02-09
影响因子：
4.8
通讯作者：
Watanabe, Hideto
中科院分区：
文献类型：
--
作者：
Sakai, Kenichiro;Kimata, Koji;Watanabe, Hideto
软骨破坏导致严重的关节疾病，如骨关节炎和脊柱疾病与背痛，软骨再生是非常低效的。软骨细胞外基质的主要组分是蛋白聚糖聚集蛋白聚糖，其含有约100个硫酸软骨素（CS）链，其赋予吸水性和抗压缩性。在这里，我们证明硫酸软骨素N-乙酰氨基半乳糖转移酶-1（CSGaINAcT-1）在软骨中CS生物合成中起着至关重要的作用。通过原位杂交和真实的时间反转录-PCR的发育软骨，CSGal-NAcT-1表现出最高水平的表达。其在软骨形成ATDC 5细胞中的表达与聚集蛋白聚糖核心蛋白的表达具有良好的相关性。在聚集蛋白聚糖转录降低的杂合子和纯合子聚集蛋白聚糖缺失软骨中，CSGal-NAcT-1转录相应地降低。软骨细胞中的酶的过表达进一步增强CS的生物合成，但不是聚集蛋白聚糖核心蛋白的生物合成，表明酶活性在细胞中不饱和，并且在过表达细胞中合成的聚集蛋白聚糖比天然分子重。通过凝胶色谱法和二糖组成分析在过表达细胞中合成的CS链，发现CS链具有相似的长度和硫酸化模式。此外，腺病毒基因传递到椎间盘中的酶显示了CS生物合成水平的大幅增加。这些观察结果表明CSGaINAcT-1过表达增加了附着于聚集蛋白聚糖核心蛋白的CS链的数量。我们的研究可能会导致一种新的治疗干预，改善软骨退行性疾病的结果。
Cartilage destruction leads to severe joint diseases, such as osteoarthritis and spinal disorders with back pain, and cartilage regeneration is very inefficient. A major component of the cartilage extracellular matrix is the proteoglycan aggrecan that contains approximately 100 chondroitin sulfate (CS) chains, which impart water absorption and resistance to compression. Here, we demonstrate that chondroitin sulfate N-acetylgalactosaminyltransferase-1 (CSGaINAcT-1) plays a critical role in CS biosynthesis in cartilage. By in situ hybridization and real time reverse transcription-PCR of developing cartilage, CSGal-NAcT-1 exhibited the highest level of expression. Its expression in chondrogenic ATDC5 cells correlated well with that of aggrecan core protein. In heterozygote and homozygote aggrecan-null cartilage where aggrecan transcription is decreased, CSGal-NAcT-1 transcription diminished accordingly. Overexpression of the enzyme in chondrocytic cells further enhanced CS biosynthesis but not that of the aggrecan core protein, indicating that the enzyme activity is not saturated in the cells and that aggrecan synthesized in the overexpressing cells is heavier than the native molecule. Analysis of the CS chains synthesized in the overexpressing cells by gel chromatography and that of disaccharide composition revealed that the CS chains had similar length and sulfation patterns. Furthermore, adenoviral gene delivery of the enzyme into intervertebral discs displayed a substantial increase in the level of CS biosynthesis. These observations indicate that CSGaINAcT-1 overexpression increases the number of CS chains attached to aggrecan core protein. Our studies may lead to a new therapeutic intervention, ameliorating the outcome of cartilage degenerative diseases.