Identification of epithelial cadherin protein forms in human sperm and evaluation of their localization in whole cells

LENTZ EM; MARÍN BRIGGILER CI; VEIGA F; VAZQUEZ-LEVIN M

Seattle, Washington, USA

Congreso; 30th Annual American Society of Andrology Meeting; 2005

American Society of Andrology

Epithelial cadherin (Ecad) belongs to a large superfamily of cell adhesion molecules showing a vast structural diversity. Ecad is a 120 KDa glycoprotein organized in five extracellular, a transmembrane, and a cytoplasmic domains. Previous studies from our group described Ecad localization in sperm and suggested its involvement in gamete interaction (Vazquez-Levin et al,  ASA 2003). The aim of the study was to perform a biochemical characterization of Ecad protein forms present in human sperm, and evaluate their localization in the whole cell. Western immunoblotting of protein sperm extracts developed with a specific antibody towards cadherin domain 5 revealed  the presence of 4 high Mr Ecad forms of 122, 105, 97 and 86 KDa. While a specific antibody towards cadherin domain 2 recognized Ecad 122, 97 and 86, an antibody towards the Ecad cytoplasmic domain detected Ecad122 and Ecad105; these results suggest that Ecad105 is truncated in the N-terminus while Ecad97 and Ecad86 are truncated in the C-teminus. Ecad86 was extracted after cell treatment with 1M NaCl, and was detected in cauda epididymal fluid and seminal plasma; moreover, its signal was diminished in protein extracts from capacitated sperm, suggesting this is a peripheral Ecad form. Sperm treatment with PLC-PI removed Ecad86 and Ecad97, indicating that Ecad97 would be anchored to the membrane by Glycosyl Phosphatidyl Inositol (GPI).  Immunocytochemistry done with the antibodies raised towards the cadherin domains 2 and 5 showed Ecad localization in the sperm head and tail. Contrasting, the antibody  towards Ecad cytoplasmic domain only stained the flagellum. In conclusion, four high Mr Ecad forms were identified in human sperm with a differential association and localization along the cell, which may be indicative of different roles in fertilization.