PDF(Pubmed)
Abstract：
Aberrant glycosylation is a hallmark of a cancer cell. One prevalent alteration is an enrichment in α2,6-linked sialylation of N-glycosylated proteins, a modification directed by the ST6GAL1 sialyltransferase. ST6GAL1 is upregulated in many malignancies including ovarian cancer. Prior studies have shown that the addition of α2,6 sialic acid to the epidermal growth factor receptor (EGFR) activates this receptor, although the mechanism was largely unknown. To investigate the role of ST6GAL1 in EGFR activation, ST6GAL1 was overexpressed in the OV4 ovarian cancer line, which lacks endogenous ST6GAL1, or knocked-down in the OVCAR-3 and OVCAR-5 ovarian cancer lines, which have robust ST6GAL1 expression. Cells with high expression of ST6GAL1 displayed increased activation of EGFR and its downstream signaling targets, AKT and NFκB. Using biochemical and microscopy approaches, including total internal reflection fluorescence microscopy, we determined that the α2,6 sialylation of EGFR promoted its dimerization and higher order oligomerization. Additionally, ST6GAL1 activity was found to modulate EGFR trafficking dynamics following EGF-induced receptor activation. Specifically, EGFR sialylation enhanced receptor recycling to the cell surface following activation while simultaneously inhibiting lysosomal degradation. 3D widefield deconvolution microscopy confirmed that in cells with high ST6GAL1 expression, EGFR exhibited greater colocalization with Rab11 recycling endosomes and reduced colocalization with LAMP1-positive lysosomes. Collectively, our findings highlight a novel mechanism by which α2,6 sialylation promotes EGFR signaling by facilitating receptor oligomerization and recycling.
摘要：
异常糖基化是癌细胞的标志。一个普遍的改变是N-糖基化蛋白的α2,6-连接唾液酸化的富集，由ST6GAL1唾液酸转移酶指导的修饰。ST6GAL1在包括卵巢癌在内的许多恶性肿瘤中上调。先前的研究表明，向表皮生长因子受体（EGFR）中添加α2,6唾液酸可激活该受体，尽管机制在很大程度上是未知的。探讨ST6GAL1在EGFR激活中的作用,ST6GAL1在OV4卵巢癌细胞系中过度表达，缺乏内源性ST6GAL1，或在OVCAR-3和OVCAR-5卵巢癌细胞系中被击倒，具有稳健的ST6GAL1表达。高表达ST6GAL1的细胞显示EGFR及其下游信号靶标的激活增加，AKT和NFκB。使用生化和显微镜方法，包括全内反射荧光（TIRF）显微镜，我们确定EGFR的α2,6唾液酸化促进其二聚化和高阶寡聚化。此外,发现ST6GAL1活性在EGF诱导的受体激活后调节EGFR运输动力学。具体来说,EGFR唾液酸化增强激活后受体向细胞表面的再循环,同时抑制溶酶体降解。3D宽场去卷积显微镜证实，在ST6GAL1高表达的细胞中，EGFR与Rab11再循环内体的共定位更大，与LAMP1阳性溶酶体的共定位减少。总的来说，我们的发现强调了α2,6唾液酸化通过促进受体寡聚化和再循环促进EGFR信号传导的新机制.
