Lectin‐gold cytochemistry of mucin oligosaccharide biosynthesis in golgi apparatus of airway secretory cells of the hamster

Abstract

To elucidate the mechanism for the biosynthesis of O-linked mucin oligosaccharides, airway secretory cells of the hamster trachea were embedded in Lowicryl K4M resin, and sections were examined by lectin-gold cytochemistry with special attention focused on the Golgi apparatus. The interrelations between the Golgi cisternae stained with five different lectins were determined by double-staining procedures using various combinations of lectins conjugated with 14-nm and 8-nm colloidal gold. Several cis cisternae were stained only with HPA (Helix pomatia agglutinin specific for terminal alpha-N-acetylgalactosamine). The next medial cisternae were not stained with HPA, but reacted positively with two lectins, GSII (Griffonia simplicifolia agglutinin II specific for terminal alpha- or beta-N-acetylglucosamine) and RCAI (Ricinus communis agglutinin I specific for beta-galactose). The trans cisternae as well as condensing and mature secretory granules were labeled with four lectins, UEAI (Ulex europaeus agglutinin I specific for terminal alpha-L-fucose) and LFA (Limax flavus agglutinin specific for terminal N-acetyl or N-glycolyl neuraminic acid) in addition to HPA and RCAI. The same number of trans cisternae were positive to HPA and UEAI, whereas LFA bound to a few transmost cisternae but fewer than were stained with HPA or UEAI. The observed sequential appearance of different sugar residues in different levels of Golgi cisternae (from cis to trans cisternae) coincides quite well with the sugar sequence of airway mucin oligosaccharide (from reducing to nonreducing ends) proposed by biochemical analysis. It is suggested that airway mucin oligosaccharides elongate during a vectorial movement through the Golgi stack from cis toward trans and that the stack consists of at least three functionally distinct segments, cis, medial, and trans; in these three segments there take place, respectively, the initial O-glycosylation of mucin core peptide, the formation of a core region of oligosaccharide chain, and the completion of chain growth by addition of terminal sugar moieties.

Cytochemical characteristics of the golgi apparatus

“…At each stack, cis, medial, and trans cisternae, as well as transmost elements (trans Golgi reticulum, trans tubular network) have been defined morphologically and biochemically. Morphological studies, successful in labeling distinct Golgi apparatus regions, include metal impregnation (Friend and Murray, 1965), enzymecytochemical and immunocytochemical localization of Golgi-associated enzymes (e.g., Dunphy et al, 1985;Ellinger and Pavelka, 1982;Hand and Oliver, 1984;Matsuo et al, 1988;Matyas and Morre, 1983;Novikoff and Novikoff, 1977;Paavola, 1978;Pavelka and Ellinger, 1983;Rambourg et al, 1988;Roth and Berger, 1982;Roth et al, 1985;Strous et al, 19831, immunocytochemical localization of various molecules traversing the Golgi stacks (e.g., Bergmann and Singer, 1983;Geuze et al, 1985;Green et al, 1981;Griffiths et al, 1989;Posthuma et al, 1984;Saraste and Kuismanen, 1984;Slot and Geuze, 1983;Yokota and Fahimi, 19811, lectinocytochemical localization of glycoconjugates (e.g., Ellinger and Pavelka, 1988;Hedman et al, 1986;Lucocq et al, 1987;Malchiodi et al, 1986;Pavelka andEllinger, 1985,1989;Quatacker, 1989;Roth, 1984;Roth et al, 1984Roth et al, ,1988Sato and Spicer, 1982;Tartakoff and Vassalli, 1983;Taatjes et al, 1988;Velasco et al, 1988;Wasano et al, 1988), and radioautographical demonstration of sug...…”

Cytochemical characterization of sulfo- and sialoglycoconjugates of human laryngeal glandular cells.

“…Glycoproteins have been reported to play protective, barrier and transport functions in the airways (Kaliner et al, 1986). Characterization of airway glycoproteins has been carried out by both biochemical and histochemical approaches (Klein et al, 1993;Lamblin et al, 1992;Sangadala et al, 1992;Castells et al, 1990Castells et al, ,1991Plotkmki et al, 1990;Wasano et al, 1988;Breg et al, 1987;Kaliner et al, 1986;Spicer, 1983,1985;Spicer et al, 1983a,b;Mazzuca et al, 1982). Sialic acid has been described as the usual terminal residue of the oligosaccharide side-chain of glycoproteins.…”

Ultrastructural localization of glycoconjugates in human bronchial glands: the subcellular organization of N- and O-linked oligosaccharide chains.

“…The ultrastructural application of colloidal gold-conjugated lectins in combination with biochemical determinations in different cells and tissues has disclosed the synthetic pathways of the glycoproteins at the ultrastructural level Wasano et al, 1988;Roth, 1984). However, this technology has not been applied to the adult human bronchial epithelium.…”