Binding of rainbow trout sperm to egg is mediated by strong carbohydrate-to-carbohydrate interaction between (KDN)GM3 (deaminated neuraminyl ganglioside) and Gg3-like epitope

^*Division of Biomembrane Research, Pacific Northwest Research Institute, Seattle, WA 98122; ^†Departments of Pathobiology and Microbiology, University of Washington, Seattle, WA 98195; ^‖Laboratory of Drug Resistance in Bacteria, Gunma University School of Medicine, Maebashi, Gunma 371-8511, Japan; and ^**Institute of Biological Chemistry, Academia Sinica, Taipei 115, Taiwan

Contributed by Sen-itiroh Hakomori

Abstract

KDNα2→3Galβ4Glcβ1Cer [(KDN)GM3] is a major (≈90%) component of total gangliosides found in sperm of rainbow trout (Oncorhynchus mykiss) and was shown to be present prominently at the sperm head by immunochemical staining with its specific mAb kdn3G. Liposomes containing (KDN)GM3 adhere specifically to GalNAcβ4Galβ4Glcβ1Cer (Gg3Cer)-coated plastic plates. Interaction between (KDN)GM3 and Gg3Cer was much stronger than that previously observed between Neu5Acα2→3Galβ4Glcβ1Cer and Gg3Cer. (KDN)GM3–Gg3Cer interaction did not require the presence of Ca²⁺ and Mg²⁺, but was enhanced in the presence of Mn²⁺. Fresh trout sperm adhered specifically to Gg3Cer-coated plates under physiological conditions, and the binding was inhibited by pretreatment of sperm with mAb kdn3G. The presence of Gg3 or Gg3-related epitope structure in the specific area surrounding the micropyle, through which sperm enter the egg, was confirmed by immunogold labeling under electron microscopy. These findings suggest that initial sperm-egg adhesion during the process of fertilization occurs when sperm adhere to the area surrounding the micropyle through specific interaction between (KDN)GM3 on the sperm head and Gg3 epitope (GalNAcβ4Galβ1→) expressed at a defined region of the egg surface membrane.

Footnotes

↵ ‡ Present address: Department of Information Systems, School of Medicine, University of Maryland, 100 North Greene Street, Room 214B, Baltimore, MD 21201.
↵ § Present address: Department of Chemical Engineering, Tokai University, Hiratsuka 259-1292, Japan.
↵ ¶ To whom reprint requests should be addressed. E-mail: hakomori{at}u.washington.edu or syinoue{at}gate.sinica.edu.tw.
↵ ‡‡ Hernaiz, M. J., Gallardo, J. A., de la Fuente, J. M., Barrientos, A. G. & Penades, S. (2001) Glycoconj. J. 18, 31 (abstr.).
Abbreviations:

BCECF-AM,

bis(carboxymethyl)carboxyfluorescein acetoxymethyl ester;

chydr,

carbohydrate;

Gb4,

GalNAcβ3Galα4Galβ4Glcβ1Cer;

Gg3Cer,

GalNAcβ4Galβ4Glcβ1Cer;

GSL,

glycosphingolipid;

KDN,

2-keto-3-deoxy-d-glycero-d-galacto-nononic acid;

(KDN)GM3,

KDNα2→3Galβ4Glcβ1Cer;

LacCer,

Galβ4Glcβ1Cer;

Lex,

Galβ4(Fucα3)GlcNAcβ1;

MF,

membranous fraction;

(Neu5Ac)GM3,

Neu5Acα2→3Galβ4Glcβ1Cer;

SRS,

salmon Ringer's solution;

VE,

vitelline envelope