Medeiros Andrea, Chiribao María Laura, Ubillos Luis, Festari María Florencia, Saldaña Jenny, Robello Carlos, Domínguez Laura, Calvete Juan José, Osinaga Eduardo
Departamento de Bioquímica, Facultad de Medicina, Universidad de la República, Montevideo, Uruguay.
Int J Parasitol. 2008 Mar;38(3-4):265-76. doi: 10.1016/j.ijpara.2007.07.015. Epub 2007 Aug 14.
Protein glycosylation is an important post-translational modification underlying host-parasite interactions, which may determine the outcome of infection. Although Mesocestoides vogae represents an important model for investigating the various aspects of cestode biology, virtually no information is available about the structure and synthesis of glycans in this parasite. In this work, focused on the initiation pathway of mucin-type O-glycosylation in M. vogae, we characterized O-glycoproteins bearing the simple mucin-type cancer-associated Tn and sialyl-Tn antigens, and the expression and activity of ppGalNAc-T, the key enzyme responsible for the first step of mucin-type O-glycosylation. Using immunohistochemistry, Tn and sialyl-Tn antigens were detected mainly in the tegument (microtriches) and in parenchymal cells. Tn expression was also observed in lateral nerve cords. Both Tn and sialyl-Tn antigens were detected in in vitro cultured parasites. Based on their electrophoretic mobility, Tn- and sialyl-Tn-bearing glycoproteins from M. vogae were separated into several components of 22 to 60 kDa. The observation that Tn and sialyl-Tn glycoproteins remained in the 0.6N perchloric acid-soluble fraction suggested that they could be good candidates for characterizing mucin-type glycosylation in this parasite. O-glycoproteins were purified and initially characterized using a proteomic approach. Immunohistochemical analysis of the tissue distribution of ppGalNAc-T revealed that this enzyme is expressed in the sub-tegumental region and in the parenchyma of the parasite. In M. vogae cultured in vitro, ppGalNAc-T was mainly detected in the suckers. Using a panel of 8 acceptor substrate synthetic peptides, we found that M. vogae ppGalNAc-T preferentially glycosylate threonine residues, the best substrates being peptides derived from human mucin MUC1 and from Trypanosoma cruzi mucin. These results suggest that M. vogae might represent a useful model to study O-glycosylation, and provide new research avenues for future studies on the glycopathobiology of helminth parasites.
蛋白质糖基化是宿主 - 寄生虫相互作用中一种重要的翻译后修饰，它可能决定感染的结果。虽然沃氏中殖孔绦虫是研究绦虫生物学各个方面的重要模型，但关于这种寄生虫中聚糖的结构和合成几乎没有可用信息。在这项工作中，我们聚焦于沃氏中殖孔绦虫粘蛋白型O - 糖基化的起始途径，对带有简单粘蛋白型癌症相关Tn和唾液酸化Tn抗原的O - 糖蛋白，以及粘蛋白型O - 糖基化第一步的关键酶ppGalNAc - T的表达和活性进行了表征。通过免疫组织化学方法，Tn和唾液酸化Tn抗原主要在皮层（微毛）和实质细胞中被检测到。在侧神经索中也观察到了Tn的表达。在体外培养的寄生虫中也检测到了Tn和唾液酸化Tn抗原。基于它们的电泳迁移率，来自沃氏中殖孔绦虫的带有Tn和唾液酸化Tn的糖蛋白被分离成了几个22至60 kDa的组分。Tn和唾液酸化Tn糖蛋白保留在0.6N高氯酸可溶部分的观察结果表明，它们可能是表征这种寄生虫中粘蛋白型糖基化的良好候选物。O - 糖蛋白被纯化并最初使用蛋白质组学方法进行了表征。对ppGalNAc - T组织分布的免疫组织化学分析表明，这种酶在寄生虫的皮层下区域和实质中表达。在体外培养的沃氏中殖孔绦虫中，ppGalNAc - T主要在吸盘处被检测到。使用一组8种受体底物合成肽，我们发现沃氏中殖孔绦虫ppGalNAc - T优先将苏氨酸残基糖基化，最佳底物是源自人粘蛋白MUC1和克氏锥虫粘蛋白的肽。这些结果表明，沃氏中殖孔绦虫可能是研究O - 糖基化的有用模型，并为未来关于蠕虫寄生虫糖病理生物学的研究提供了新的研究途径。
