Mucin-type O-glycosylation in Fasciola hepatica:: characterisation of carcinoma-associated Tn and sialyl-Tn antigens and evaluation of UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferase activity
Mucin-type O-glycosylation in Fasciola hepatica:: characterisation of carcinoma-associated Tn and sialyl-Tn antigens and evaluation of UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferase activity
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DOI：
10.1016/s0020-7519(02)00231-x
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发表时间：
2003-01-01
影响因子：
4
通讯作者：
Osinaga, E
中科院分区：
文献类型：
--
作者：
Freire, T;Casaravilla, C;Osinaga, E
简单的粘蛋白型癌症相关O-聚糖结构，如Tn抗原（GalNAc-0-Ser/Thr），由某些蠕虫寄生虫表达。这些抗原参与几种类型的受体-配体相互作用，并且它们是免疫治疗的潜在靶点。本工作的目的是研究肝片吸虫粘蛋白型O-糖基化的起始途径，对Tn和唾液酸-Tn抗原进行生化和免疫组织化学表征，并评估催化O-聚糖生物合成第一步的ppGaNTase活性。使用ELISA，Tn和唾液酸-Tn抗原检测主要在体细胞和脱氧胆酸盐提取物。免疫荧光分析显示，Tn抗原优先表达在睾丸，而唾液酸-Tn糖蛋白分布更广泛，存在于实质细胞，基底膜的被膜，和顶端表面的上皮细胞内衬盲肠。根据其电泳迁移率，Tn糖蛋白被解析为10，37，76，125，170和205 kDa的6个组分，唾液酸Tn组分显示出28和32 kDa的表观分子量，以及90-110和170-190 kDa的两条宽带。只有76 kDa的Tn-糖蛋白仍然在0.6 N高氯酸可溶性馏分的观察表明，它可能是一个很好的候选人在这种寄生虫的粘蛋白表征。ppGaNTase在pH 7-7.5和37 ℃时活性最高，表明Mn ~（2+）是最佳的二价阳离子激活剂。使用一组九个合成肽作为受体底物，我们发现F。Hepatica ppGaNTase能够糖基化苏氨酸和丝氨酸，最好的底物是来源于人粘蛋白（MUC 2和MUC 6）的串联重复区以及克氏锥虫和布氏锥虫糖蛋白的肽。本文报道的结果构成了F. hepatica，并可能有助于确定这种寄生虫的新的生物学特性以及宿主-寄生虫关系。(C)2002年澳大利亚寄生虫学会由爱思唯尔科技有限公司出版。保留所有权利。
Simple mucin-type cancer-associated O-glycan structures, such as the Tn antigen (GalNAc-O-Ser/Thr), are expressed by certain helminth parasites. These antigens are involved in several types of receptor-ligand interactions, and they are potential targets for immunotherapy. The aim of this work was to study the initiation pathway of mucin-type O-glycosylation in Fasciola hepatica, performing a biochemical and immunohistochemical characterisation of Tn and sialyl-Tn antigens, and evaluating the ppGaNTase activity, which catalyses the first step in O-glycan biosynthesis. Using ELISA, both Tn and sialyl-Tn antigens were detected predominantly in the somatic and deoxycholate extracts. Immunofluorescence analysis revealed that Tn antigen is preferentially expressed in testis, while sialyl-Tn glycoproteins were more widely distributed, being present in parenchymal cells, basal membrane of the tegument, and apical surface of epithelial cells lining the caeca. On the basis of their electrophoretic mobility, Tn glycoproteins were resolved as six components of 10, 37, 76, 125, 170 and 205 kDa, and sialyl-Tn components showed an apparent molecular mass of 28 and 32 kDa, and two broad bands of 90-110 and 170-190 kDa. The observation that only the 76 kDa Tn-glycoprotein remained in the 0.6 N perchloric acid-soluble fraction suggests that it could be a good candidate for mucin characterisation in this parasite. The ppGaNTase activity showed its maximal activity at pH 7-7.5 and 37 degreesC, showing that Mn2+ was the best divalent cation activator. Using a panel of nine synthetic peptides as acceptor substrates, we found that F. hepatica ppGaNTase was able to glycosylate both threonines and serines, the best substrates being the peptides derived from the tandem repeat region of human mucins (MUC2 and MUC6), and from Trypanosoma cruzi and Trypanosoma brucei glycoproteins. The results reported here constitute the first evidence on O-glycosylation pathways in F. hepatica, and may help to identify new biological characteristics of this parasite as well as of the host-parasite relationship. (C) 2002 Australian Society for Parasitology Inc. Published by Elsevier Science Ltd. All rights reserved.