Cytochemical characterization of oligosaccharide side chains of the glycoproteins of rat zona pellucida: an ultrastructural study.
BACKGROUND
The zona pellucida (ZP), an extracellular matrix which surrounds mammalian oocytes, is formed by different glycoproteins. Several studies have revealed that carbohydrate residues present in glycoproteins of ZP play a key role in the sperm-egg recognition. However, the origin and the biochemical composition of ZP remain to be completely resolved.


METHODS
ZP glycoproteins from rat ovarian follicles were investigated at light and electron microscopic level by the application of lectins conjugated to peroxidase, digoxigenin, and colloidal gold in combination with enzyme and chemical treatment. A quantitative analysis was also performed.


RESULTS
ZP shows reactivity to WGA, DSA, LFA, AAA, RCA I, and MAA. SBA and PNA showed a variable reactivity ranging from negative to strongly positive. A uniform pattern of binding throughout ZP was observed with DSA, Con A, AAA, MAA, and LFA. However, labeling by RCA I and SBA was higher in the outer ZP while PNA and WGA showed a higher binding in the inner ZP. Lectin reactivity was detected in cortical granules, endoplasmic reticulum, Golgi apparatus, vesicles, and multivesicular bodies of oocytes.


CONCLUSIONS
ZP contained the terminal disaccharides Gal beta 1,4GlcNAc, Gal beta 1,3GalNAc, and GalNAc beta 1,3Gal and the trisaccharides Neu5Ac alpha 2, 3Gal beta 1,4GlcNAc, Neu5Ac-Gal beta 1,3GalNAc, and Neu5Ac-GalNAc beta 1,3Gal sequences. The occurrence of Fucose residues alpha 1,6 linked to the inner core region of N-linked glycoproteins of ZP was demonstrated by the use of several fucose-specific lectins. Methylation-saponification treatment in combination with lectin cytochemistry reveals that Gal, GalNAc, and polyllactosamine residues of rat ZP glycoproteins contain sulphated groups. The reactivity observed in ooplasmic vesicles was similar to that of ZP, thus suggesting that the oocyte is the site of synthesis of ZP glycoproteins.
Avilés
M
M
Department of Cell Biology, School of Medicine, University of Murcia, Spain.
Martínez-Menárguez
J A
JA
Castells
M T
MT
Madrid
J F
JF
Ballesta
J
J
D000818:Animals; D005260:Female; D006023:Glycoproteins / Q000737:chemistry*; D006651:Histocytochemistry; D037102:Lectins; D008854:Microscopy, Electron; D009439:Neuraminidase / Q000494:pharmacology; D009844:Oligosaccharides / Q000737:chemistry*; D009865:Oocytes / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D014018:Tissue Distribution; D015044:Zona Pellucida / Q000378:metabolism* / Q000648:ultrastructure
D006023:Glycoproteins; D037102:Lectins; D009844:Oligosaccharides; D009439:Neuraminidase