Uyama Toru, Ishida Miho, Izumikawa Tomomi, Trybala Edward, Tufaro Frank, Bergström Tomas, Sugahara Kazuyuki, Kitagawa Hiroshi
Department of Biochemistry, Kobe Pharmaceutical University, 4-19-1 Motoyamakita-machi, Higashinada-ku, Kobe 658-8558, Japan.
J Biol Chem. 2006 Dec 15;281(50):38668-74. doi: 10.1074/jbc.M609320200. Epub 2006 Oct 13.
We have demonstrated a defect in expression of chondroitin 4-O-sulfotransferase-1 (C4ST-1) in murine sog9 cells, which are poorly sensitive to infection by herpes simplex virus type 1 (HSV-1). Sog9 cells were previously isolated as CS-deficient cells from gro2C cells, which were partially resistant to HSV-1 infection and defective in the expression of heparan sulfate (HS) because of a splice site mutation in the EXT1 gene encoding the HS-synthesizing enzyme. Here we detected a small amount of CS chains in sog9 cells with a drastic decrease in 4-O-sulfation compared with the parental gro2C cells. RT-PCR revealed that sog9 cells had a defect in the expression of C4ST-1 in addition to EXT1. Gel filtration analysis showed that the decrease in the amount of CS in sog9 cells was the result of a reduction in the length of CS chains. Transfer of C4ST-1 cDNA into sog9 cells (sog9-C4ST-1) restored 4-O-sulfation and amount of CS, verifying that sog9 cells had a specific defect in C4ST-1. Furthermore, the expression of C4ST-1 rendered sog9 cells significantly more susceptible to HSV-1 infection, suggesting that CS modified by C4ST-1 is sufficient for the binding and infectivity of HSV-1. Analysis of CS chains of gro2C and sog9-C4ST-1 cells revealed a considerable proportion of the E disaccharide unit, consistent with our recent finding that this unit is an essential component of the HSV receptor. These results suggest that C4ST-1 regulates the expression of the E disaccharide unit and the length of CS chains, the features that facilitate infection of cells by HSV-1.
我们已经证明，在对1型单纯疱疹病毒（HSV-1）感染敏感性较差的小鼠sog9细胞中，硫酸软骨素4-O-磺基转移酶-1（C4ST-1）的表达存在缺陷。Sog9细胞先前是从gro2C细胞中分离出来的硫酸软骨素（CS）缺陷细胞，gro2C细胞对HSV-1感染具有部分抗性，并且由于编码硫酸乙酰肝素（HS）合成酶的EXT1基因中的剪接位点突变，其硫酸乙酰肝素（HS）表达存在缺陷。在这里，我们在sog9细胞中检测到少量的CS链，与亲代gro2C细胞相比，其4-O-硫酸化程度急剧下降。逆转录聚合酶链反应（RT-PCR）显示，除EXT1外，sog9细胞中C4ST-1的表达也存在缺陷。凝胶过滤分析表明，sog9细胞中CS量的减少是CS链长度缩短的结果。将C4ST-1 cDNA导入sog9细胞（sog9-C4ST-1）可恢复4-O-硫酸化和CS量，证实sog9细胞在C4ST-1方面存在特异性缺陷。此外，C4ST-1的表达使sog9细胞对HSV-1感染的敏感性显著增加，这表明经C4ST-1修饰的CS足以支持HSV-1的结合和感染性。对gro2C和sog9-C4ST-1细胞的CS链分析揭示了相当比例的E二糖单元，这与我们最近的发现一致，即该单元是HSV受体的重要组成部分。这些结果表明，C4ST-1调节E二糖单元的表达和CS链的长度，这些特征有利于HSV-1感染细胞。
