PARENT SESSION
PLATFORM SESSION 19: SPERM-EGG BINDING AND EGG ACTIVATION
Chair: Miller, David¹, ¹
Co-chair: Rivera, Rocio¹, ¹
Harborside E
2:00 PM-4:00 PM

(432) PIG SPERM RAFTS HAVE ZONA PELLUCIDA-BINDING ABILITY AND CONTAIN MAMMALIAN MALE GERM CELL-SPECIFIC SULFOGALACTOSYLGLYCEROLIPID (SGG) AND ARYLSULFATASE-A (AS-A).

Bou Khalil, Maroun^1,2, Chakrabandhu, Krittalak¹, Carmona, Euridice¹, Xu, Hongbin¹, Vuong, Ngoc^1,2, Kumarathasan, Premkumari³, de Silva, Solange¹, Carrier, Danielle², Wong, Patrick², Tanphaichitr, Nongnuj^1,2,4, ¹ Hormones, Growth, and Development Research Group, Ottawa, CA² Department of Biochemistry, Microbiology and Immunology, Ottawa, CA³ Growth and Development Section, Environmental Health Sciences Bureau, Ottawa, CA⁴ Human In Vitro Fertilization Program, Ottawa, CA

ABSTRACT- We have previously shown that sulfogalactosylglycerolipid (SGG) and its binding protein, arylsulfatase A (AS-A), exist selectively on the male germ cell plasma membrane and play important roles in mammalian sperm-egg binding. In addition, SGG was also shown to interact with cholesterol and saturated phospholipids by Fourier transform infrared (FTIR) spectroscopy. The interaction between SGG and cholesterol in the lipid complex confers insolubility in Triton X-100, an inherent property of raft membranes. Significantly, rafts from capacitated and non-capacitated pig sperm floated, due to their high lipid content, as a light scattering band, to 5-7% sucrose following sucrose-gradient ultracentrifugation of the TritonX-100 sperm extract. High performance thin layer chromatography (HPTLC) indicated that SGG was a major lipid component in these rafts. In fact, ~60% and 75% of total SGG in non-capacitated and capacitated sperm, respectively, was in the isolated rafts. Gas chromatography-mass spectrometry (GC-MS) analysis of raft phosphatidylcholine (PC) and phosphatidylethanolamine (PE) revealed that they contained mainly saturated hydrocarbon chains (16:0 and 18:0). Immunoblotting revealed the presence of SGG binding proteins (AS-A and Hsp70), zona-pellucida (ZP) adhesive molecules (spermadhesin and -galactosyltransferase), signaling proteins (adenylyl cyclase and G_i), and tyrosine phosphorylated 66- and 59-kDa proteins. These isolated sperm rafts also have ZP binding ability, as shown by the micro-titer plate binding assay. Isolated sperm rafts are therefore representative physiological model membranes for studying the roles of SGG, AS-A, and other ZP adhesive molecules in ZP binding.

KEY WORDS: Rafts, Zona-Pellucida Binding, sulfogalactosylglycerolipid , Arylsulfatase A