Division of Biomembrane Research, Pacific Northwest Research Institute, Seattle, WA 98122, USA.
Proc Natl Acad Sci U S A. 2011 Oct 25;108(43):17690-5. doi: 10.1073/pnas.1115191108. Epub 2011 Oct 17.
The process termed "epithelial-mesenchymal transition" (EMT) was originally discovered in ontogenic development, and has been shown to be one of the key steps in tumor cell progression and metastasis. Recently, we showed that the expression of some glycosphingolipids (GSLs) is down-regulated during EMT in human and mouse cell lines. Here, we demonstrate the involvement of GalNAc-type (or mucin-type) O-glycosylation in EMT process, induced with transforming growth factor β (TGF-β) in human prostate epithelial cell lines. We found that: (i) TGF-β treatment caused up-regulation of oncofetal fibronectin (onfFN), which is defined by mAb FDC6, and expressed in cancer or fetal cells/tissues, but not in normal adult cells/tissues. The reactivity of mAb FDC6 requires the addition of an O-glycan at a specific threonine, inside the type III homology connective segment (IIICS) domain of FN. (ii) This change is associated with typical EMT characteristics; i.e., change from epithelial to fibroblastic morphology, enhanced cell motility, decreased expression of a typical epithelial cell marker, E-cadherin, and enhanced expression of mesenchymal markers. (iii) TGF-β treatment up-regulated mRNA level of FN containing the IIICS domain and GalNAc-T activity for the IIICS domain peptide substrate containing the FDC6 onfFN epitope. (iv) Knockdown of GalNAc-T6 and T3 inhibited TGF-β-induced up-regulation of onfFN and EMT process. (v) Involvement of GSLs was not detectable with the EMT process in these cell lines. These findings indicate the important functional role of expression of onfFN, defined by site-specific O-glycosylation at IIICS domain, in the EMT process.
被称为“上皮-间充质转化”（EMT）的过程最初是在个体发生发育中发现的，并且已被证明是肿瘤细胞进展和转移的关键步骤之一。最近，我们表明在人源和鼠源细胞系中 EMT 过程中某些糖脂（GSL）的表达下调。在这里，我们证明了 GalNAc 型（或粘蛋白型）O-糖基化在 TGF-β诱导的人前列腺上皮细胞系 EMT 过程中的参与。我们发现：（i）TGF-β处理导致肿瘤衍生型纤维连接蛋白（onfFN）的上调，该蛋白由 mAb FDC6 定义，并在癌症或胎儿细胞/组织中表达，但在正常成年细胞/组织中不表达。mAb FDC6 的反应性需要在 FN 的 III 型同源连接区（IIICS）域内的特定苏氨酸处添加 O-聚糖。（ii）这种变化与典型的 EMT 特征相关；即，从上皮形态转变为成纤维细胞形态，增强细胞迁移能力，降低典型上皮细胞标志物 E-钙粘蛋白的表达，增强间充质标志物的表达。（iii）TGF-β处理上调了含有 IIICS 结构域的 FN 的 mRNA 水平，以及 GalNAc-T 活性，用于含有 mAb FDC6 onfFN 表位的 IIICS 结构域肽底物。（iv）GalNAc-T6 和 T3 的敲低抑制了 TGF-β诱导的 onfFN 上调和 EMT 过程。（v）在这些细胞系中，EMT 过程中未检测到 GSL 的参与。这些发现表明，在 EMT 过程中，特定位点 O-糖基化定义的 onfFN 的表达具有重要的功能作用。
