Gao Wei, Xu Yongmei, Liu Jian, Ho Mitchell
Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Key Laboratory of Human Functional Genomics of Jiangsu Province, Department of Cell Biology, School of Basic Medical Science, Nanjing Medical University, Nanjing, Jiangsu, 210029, China.
Sci Rep. 2016 May 17;6:26245. doi: 10.1038/srep26245.
Heparan sulfate (HS) is a polysaccharide known to modulate many important biological processes, including Wnt signaling. However, the biochemical interaction between HS and Wnt molecules is not well characterized largely due to the lack of suitable methods. To determine the Wnt binding domain in HS, we used a Wnt signaling-inhibitory antibody (HS20) and a panel of synthetic HS oligosaccharides with distinct lengths and sulfation modifications. We found that the binding of HS20 to heparan sulfate required sulfation at both the C2 position (2-O-sulfation) and C6 position (6-O-sulfation). The oligosaccharides with the greatest competitive effect for HS20 binding were between six and eight saccharide residues in length. Additionally, a four residue-long oligosaccharide could also be recognized by HS20 if an additional 3-O-sulfation modification was present. Furthermore, similar oligosaccharides with 2-O, 6-O and 3-O-sulfations showed inhibition for Wnt activation. These results have revealed that HS20 and Wnt recognize a HS structure containing IdoA2S and GlcNS6S, and that the 3-O-sulfation in GlcNS6S3S significantly enhances the binding of both HS20 and Wnt. This study provides the evidence for identifying the Wnt binding domain in HS and suggests a therapeutic approach to target the interaction of Wnt and HS in cancer and other diseases.
硫酸乙酰肝素（HS）是一种已知可调节许多重要生物过程的多糖，包括Wnt信号传导。然而，由于缺乏合适的方法，HS与Wnt分子之间的生化相互作用尚未得到很好的表征。为了确定HS中的Wnt结合结构域，我们使用了一种Wnt信号抑制抗体（HS20）和一组具有不同长度和硫酸化修饰的合成HS寡糖。我们发现HS20与硫酸乙酰肝素的结合需要在C2位置（2-O-硫酸化）和C6位置（6-O-硫酸化）都进行硫酸化。对HS20结合具有最大竞争效应的寡糖长度在6至8个糖残基之间。此外，如果存在额外的3-O-硫酸化修饰，一个四残基长的寡糖也可以被HS20识别。此外，具有2-O、6-O和3-O-硫酸化的类似寡糖对Wnt激活有抑制作用。这些结果表明，HS20和Wnt识别含有艾杜糖醛酸2-硫酸酯（IdoA2S）和N-乙酰葡糖胺6-硫酸酯（GlcNS6S）的HS结构，并且GlcNS6S3S中的3-O-硫酸化显著增强了HS20和Wnt的结合。这项研究为鉴定HS中的Wnt结合结构域提供了证据，并提出了一种针对癌症和其他疾病中Wnt与HS相互作用的治疗方法。
