Structures and developmental alterations of N-glycans of zebrafish embryos

Zebrafish is a model organism suitable for studying vertebrate development. We analyzed the N-glycan structures of zebrafish embryos and their alterations during zebrafish embryogenesis to obtain basic data for studying the roles of N-glycosylation. Multiple modes of high-performance liquid chromatography and multistage mass spectrometry were used for structural analysis of N-glycans. The N-glycans from deyolked embryos at 36 hours postfertilization, a mid-pharyngula stage, contained relatively higher amounts of complex-and hybrid-type glycans with LacNAc (Gal beta 1-4GlcNAc) and/or sialyl LacNAc without additional beta 1,4-Gal, which are commonly found in mammalian tissues, as well as abundant oligomannose-type glycans. Some of the complex-and hybrid-type glycans possessed various extended LacNAc structures, such as Gal beta 1-4LacNAc, LacNAc-repeat or unique (+/- dHex)-GalNAc alpha 1-GlcNAc beta 1-LacNAc. In contrast, the yolk of the embryo contains predominant oligomannose-type glycans and complex-type glycans with Gal beta 1-4(Siaa2-3) Gal beta 1-4(Fuca1-3) GlcNAc antennae. N-Glycan profiles obtained from deyolked embryos at different stages showed stage-dependent variation of complex-and hybrid-type glycans. At gastrula and early segmentation stages, complex-and hybrid-type glycans were minor components, and their antenna structures were mainly sialyl LacdiNAc (Sia alpha 2-6GalNAc beta 1-4GlcNAc). From the mid-segmentation to pharyngula stages, those with LacNAc and/or alpha 2,6-sialyl LacNAc antenna structures increased remarkably, and those with alpha 2,3-sialyl LacNAc antenna, core alpha 1,6-Fuc and bisecting GlcNAc modifications increased gradually. These results suggest the presence of mechanisms for regulating the antenna structures of complex/ hybrid N-glycan biosynthesis in the phylotypic stage of vertebrate development.