Role of glycosaminoglycan-synthesizing enzymes in corneal transparency

(2)N-乙酰氨基葡萄糖6-磺酸转移酶(GlcNAc6ST)：GlcNAc6ST参与角蛋白硫酸酯链上GlcNAc残基的硫酸盐化。该酶可作为角膜分化的标志物。该基因开放阅读框序列的大部分(90%)已被测定。该序列与人GlcNAc6ST-1有83.0%的同源性，但与人角膜特异性GlcNAc6ST-5不同。此外，该序列5‘端300bps的序列与GlcNAc6ST-1的同源性很低。(3)软骨素合成酶和β-葡萄糖转移酶(Gal-T)：软骨素合成酶参与软骨素链的延长，Gal-T参与角蛋白硫酸酯链的延长。两种酶均可作为角膜脱分化或转分化的标志。这两个cDNA的开放阅读框序列已经确定。鸡软骨素合酶基因的序列与人软骨素合酶-1的同源性为82.0%，而鸡Gal-T与已报道的鸡Gal-T的同源性为82.0%。用实时荧光定量聚合酶链式反应检测培养的基质细胞中这两种酶的表达水平。在传代培养(第三代培养)过程中，两种酶的水平变化不大。

cDNAs of some glycosyltransferases and sulfotransferases which are involved in biosynthesis of glycosaminoglycan chain were cloned from chick corneal cells.(2)N-Acetylglucosamine 6-sulfotransferase (GlcNAc6ST) : GlcNAc6ST is involved in a sulfation of GlcNAc residue of keratan sulfate chain. This enzyme could be a marker for the corneal differentiation. Most part (90%) of the sequence of ORF of the cDNA has been determined. The sequence showed 83.0% homology with human GlcNAc6ST-1, but not with human cornea-specific GlcNAc6ST-5. Furthermore, 300 bps of 5'-region of the sequence showed very low homology even with the GlcNAc6ST-1.(3)Chondroitin synthasae and β-glucosyltransferases (Gal-T) : Chondroitin synthase is involved in the elongation of chondroitin chain and Gal-T in the elongation of keratan sulfate chain. Both enzymes could be marker for the dedifferentiation or transdifferentiation from cornea. The sequences of ORFs of both cDNAs have been determined. The sequence of chick chondroitin synthase cDNA showed 82.0% homology with human chondroitin synthase-1, while that of chick Gal-T was identical with chick Gal-T I reported previously. Expression levels of these two enzymes in cultured stromal cells were examined by real-time PCR method. The levels of the two enzymes did not changed so much during the passage of culture (tertiary culture).