Cholera intoxication of human enteroids reveals interplay between decoy and functional glycoconjugate ligands.
Cholera intoxication of human enteroids reveals interplay between decoy and functional glycoconjugate ligands.
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DOI：
10.1093/glycob/cwad069
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发表时间：
2023-10-30
期刊：
影响因子：
4.3
通讯作者：
中科院分区：
文献类型：
--
作者：
先前对霍乱毒素（CT）结合和中毒的研究依赖于人类结肠癌衍生的上皮细胞。虽然这些转化的细胞系是有益的，但它们既不源自发生中毒的小肠，也不代表小肠上皮细胞（SI-EC）的多样性和个体之间糖缀合物表达的变化。在这里，我们使用来自多个供体的空肠活检的人肠类物质来研究人未转化的 SI-EC 的 CT 结合和中毒。我们调节鞘糖脂、糖蛋白和特定聚糖的表面表达，以区分每种聚糖/糖缀合物的作用。霍乱毒素 B 亚基 (CTB) 突变体的产生是为了破译每种糖缀合物对不同结合位点的偏好以及 CT 结合与中毒之间的相关性。人肠类含有微量的 GM1，但其他鞘糖脂可能会导致 CT 中毒。我们发现，抑制岩藻糖基化或 O-糖基化会使肠类对 CT 中毒敏感。这可能是由于去除了与 CTB 的非规范位点结合的岩藻糖基化“诱饵样配体”和/或与规范位点结合的 Gal/GalNAc 末端糖缀合物的可用性增加的结果。此外，同时抑制岩藻糖基化和 O-糖基化增加了额外 Gal/GalNAc 末端糖缀合物的可用性，但抵消了由于岩藻糖减少而抑制 O-糖基化引起的 CT 中毒致敏作用。这意味着岩藻糖具有作为功能性聚糖和诱饵的双重作用，其相互作用影响 CT 结合和中毒。最后，虽然来自不同捐赠者的肠类的结果相似，但它们并不相同，这表明人类遗传变异在决定 CT 敏感性方面发挥了作用。
Prior research on cholera toxin (CT) binding and intoxication has relied on human colonic cancer derived epithelial cells. While these transformed cell lines have been beneficial, they neither derive from small intestine where intoxication occurs, nor represent the diversity of small intestinal epithelial cells (SI-ECs) and variation in glycoconjugate expression among individuals. Here, we used human enteroids, derived from jejunal biopsies of multipledonors to study CT binding and intoxication of human non-transformed SI-ECs. We modulated surface expression of glycosphingolipids, glycoproteins and specific glycans to distinguish the role of each glycan/glycoconjugate. Cholera-toxin-subunit-B (CTB) mutants were generated to decipher the preference of each glycoconjugate to different binding sites and the correlation between CT binding and intoxication. Human enteroids contain trace amounts of GM1, but other glycosphingolipids may be contributing to CT intoxication. We discovered that inhibition of either fucosylation or O-glycosylation sensitize enteroids to CT-intoxication. This can either be a consequence of the removal of fucosylated “decoy-like-ligands” binding to CTB’s non-canonical site and/or increase in the availability of Gal/GalNAc-terminating glycoconjugates binding to the canonical site. Furthermore, simultaneous inhibition of fucosylation and O-glycosylation increased the availability of additional Gal/GalNAc-terminating glycoconjugates but counteracted the sensitization in CT intoxication caused by inhibiting O-glycosylation because of reduction in fucose. This implies a dual role of fucose as a functional glycan and a decoy, the interplay of which influences CT binding and intoxication. Finally, while the results were similar for enteroids from different donors, they were not identical, pointing to a role for human genetic variation in determining sensitivity to CT.
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影响因子：
16.6
作者：
Mandal, Pintu K.;Branson, Thomas R.;Hayes, Edward D.;Ross, James F.;Gavin, Jose A.;Daranas, Antonio H.;Turnbull, W. Bruce
通讯作者：
Turnbull, W. Bruce
影响因子：
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影响因子：
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作者：
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Donowitz M
影响因子：
3.8
作者：
Fakhari, Amir;Baoum, Abdulgader;Siahaan, Teruna J.;Le, Khoi Ba;Berkland, Cory
通讯作者：
Berkland, Cory