Identification of mannose moieties in N‐ and O‐linked oligosaccharides of the primordial germ cells of Xenopus embryos

Abstract

The presence of mannose (Man) in the glycoconjugates of primordial germ cells (PGCs) of Xenopus embryos was elucidated by lectin histochemistry with Concanavalin A (Con A) and snowdrop (Galanthus nivalis) bulb lectin (GNA), in combination with deglycosylative pretreatments: beta-elimination, which removes O-linked oligosaccharides, and incubation with Peptide N glycosidase F (PNGase F), which removes N-linked glycan chains. In addition, histochemistry with Con A, which binds to Man and glucose (Glc), was also performed after glucose-oxidase incubation, which converts Glc into gluconic acid, and GNA was carried out after acid hydrolysis, which removes terminal sialic acid (NeuAc) moieties. PGCs were analyzed during their migration over the mesentery until the genital ridge, and after colonization of this gonad anlage. The results showed that for both lectins: (1) the PGCs and other surrounding tissue showed a similar binding pattern, and (2) the staining in the PGCs was similar in the developmental stages studied. Labeling with Con A was due to Man, and not to Glc, as shown after incubation with glucose-oxidase, and it was assumed that Man was in N-linked oligosaccharides. However, GNA labeling was mainly due to O-linked oligosaccharides, because the pretreatment of beta-elimination turned cells negative. Moreover, acid hydrolysis pretreatment gave rise to a stronger GNA-staining, suggesting that either Man was also in subterminal position to NeuAc or some Man-containing glycans were unmasked after removal of NeuAc from other oligosaccharide chains.