Ben-Zaken Olga, Tzaban Salit, Tal Yuval, Horonchik Lior, Esko Jeffrey D, Vlodavsky Israel, Taraboulos Albert
Department of Oncology, Hadassah-University Hospital, Jerusalem 91120, Israel.
J Biol Chem. 2003 Oct 10;278(41):40041-9. doi: 10.1074/jbc.M301152200. Epub 2003 Jul 18.
During prion diseases, the host protein PrPC is refolded into an abnormal conformer "prion" PrPSc. Histological and pharmacological data have suggested that glycosaminoglycans may be involved in the development of prion diseases. Here we present the first direct evidence that cellular glycosaminoglycans play a role in the biogenesis of PrPSc in prion-infected ScN2a cells. When ScN2a cells were incubated with estradiol beta-d-xyloside to inhibit the glycosylation of proteoglycans, PrPSc was vastly reduced. Treating ScN2a-M cells with heparinase III, but not with heparinase I or chondroitinase ABC, caused a profound reduction of PrPSc. In contrast, neither the amount of PrPC nor its subcellular distribution were affected as assayed by immunofluorescence microscopy and flotation procedures. In vitro treatment of ScN2a membranes with heparinase III at either neutral or acidic pH did not reduce the level of protease-resistant PrPSc. The inhibitor of sulfation, sodium chlorate, vastly reduces PrPSc in ScN2a cells (Gabizon, R., Meiner, Z., Halimi, M., and Ben-Sasson, S. A. (1993) J. Cell. Physiol. 157, 319-325). Both soluble heparan sulfate and chondroitin sulfate partially restored the level of PrPSc in chlorate-treated cells. We conclude that heparinase III-sensitive, presumably undersulfated, cellular heparan sulfate plays a significant role in the biogenesis of PrPSc in ScN2a cells.
在朊病毒疾病期间,宿主蛋白PrPC会重新折叠成异常构象体“朊病毒”PrPSc。组织学和药理学数据表明,糖胺聚糖可能参与了朊病毒疾病的发展。在此,我们提供了首个直接证据,证明细胞糖胺聚糖在朊病毒感染的ScN2a细胞中PrPSc的生物合成过程中发挥作用。当ScN2a细胞与β-D-木糖雌二醇一起孵育以抑制蛋白聚糖的糖基化时,PrPSc大幅减少。用肝素酶III处理ScN2a-M细胞,而非肝素酶I或软骨素酶ABC,会导致PrPSc显著减少。相比之下,通过免疫荧光显微镜和浮选程序检测发现,PrPC的量及其亚细胞分布均未受到影响。在中性或酸性pH条件下,用肝素酶III对ScN2a细胞膜进行体外处理,并不会降低蛋白酶抗性PrPSc的水平。硫酸化抑制剂氯酸钠可大幅降低ScN2a细胞中的PrPSc(Gabizon, R., Meiner, Z., Halimi, M., and Ben-Sasson, S. A. (1993) J. Cell. Physiol. 157, 319 - 325)。可溶性硫酸乙酰肝素和硫酸软骨素均可部分恢复氯酸钠处理细胞中PrPSc的水平。我们得出结论,对肝素酶III敏感、可能硫酸化不足的细胞硫酸乙酰肝素在ScN2a细胞中PrPSc的生物合成过程中发挥着重要作用。