Visualizing enveloping layer glycans during zebrafish early embryogenesis
Visualizing enveloping layer glycans during zebrafish early embryogenesis
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DOI：
10.1073/pnas.0912081107
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发表时间：
2010-06-08
影响因子：
11.1
通讯作者：
Bertozzi, Carolyn R.
中科院分区：
文献类型：
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作者：
Baskin, Jeremy M.;Dehnert, Karen W.;Bertozzi, Carolyn R.
通过使用非侵入性成像技术，可以在细胞和分子水平上监测发育事件。在可能作为成像分析目标的生物分子中，聚糖由于其在细胞表面的主要位置而占据优势地位。我们之前描述了一种在斑马鱼幼体发育过程中对聚糖进行成像的化学方法；然而，我们无法在胚胎发生的最初24小时（这是发育中非常活跃的时期）检测到聚糖。在这里，我们报告了一种对聚糖进行成像的方法，该方法能够在斑马鱼胚胎发生的早期阶段使聚糖在包被层中可视化。我们在单细胞阶段将叠氮糖微量注射到胚胎中，让斑马鱼发育，然后用无铜点击化学方法检测代谢标记的聚糖。黏蛋白型O -聚糖早在受精后7小时（发育的原肠胚阶段）就可以成像。此外，我们使用一种非代谢方法用一种独立的化学方法标记唾液酸化聚糖，从而能够同时对这两种不同类型的聚糖进行成像。对聚糖运输的成像分析揭示了聚糖在秒级时间尺度上的剧烈重组，包括向有丝分裂细胞的分裂沟快速迁移。这些研究深入了解了胚胎发生过程中包被层中聚糖的生物合成和动态，并为通过微量注射适当功能化的生物合成前体对其他生物分子靶点进行成像提供了一个平台。
Developmental events can be monitored at the cellular and molecular levels by using noninvasive imaging techniques. Among the biomolecules that might be targeted for imaging analysis, glycans occupy a privileged position by virtue of their primary location on the cell surface. We previously described a chemical method to image glycans during zebrafish larval development; however, we were unable to detect glycans during the first 24 hours of embryogenesis, a very dynamic period in development. Here we report an approach to the imaging of glycans that enables their visualization in the enveloping layer during the early stages of zebrafish embryogenesis. We microinjected embryos with azidosugars at the one-cell stage, allowed the zebrafish to develop, and detected the metabolically labeled glycans with copper-free click chemistry. Mucin-type O-glycans could be imaged as early as 7 hours post-fertilization, during the gastrula stage of development. Additionally, we used a nonmetabolic approach to label sialylated glycans with an independent chemistry, enabling the simultaneous imaging of these two distinct classes of glycans. Imaging analysis of glycan trafficking revealed dramatic reorganization of glycans on the second time scale, including rapid migration to the cleavage furrow of mitotic cells. These studies yield insight into the biosynthesis and dynamics of glycans in the enveloping layer during embryogenesis and provide a platform for imaging other biomolecular targets by microinjection of appropriately functionalized biosynthetic precursors.