Synthesis and glycosylation of CD52, the major 'maturation-associated' antigen of rat spermatozoa, in the cauda epididymidis.
A western and lectin blot analysis was performed of the major 'maturation-associated' antigen of rat spermatozoa, which is the rat counterpart of human CD52. In the absence of a suitable antibody, direct study of this approximately 26 kDa antigen, named previously SMemG, had been difficult. In the present study, these problems were overcome by raising a polyclonal antibody against a chemosynthetic peptide predicted from the cDNA sequence of the antigen. The antibody bound to a glycoprotein of rat cauda epididymidal tissue and spermatozoa, this glycoprotein was cleaved by phosphatidylinositol-specific phospholipase C and, after deglycosylation, was reduced to approximately 6 kDa. Northern blot analysis confirmed that the CD52 mRNA was transcribed only post-testicularly, and antibody binding to testicular and sperm proteins of different molecular masses was shown to be nonspecific. Flow cytometry also indicated that the antigen was inserted into the sperm membrane during epididymal transit. Moreover, despite the presence of CD52 mRNA in all parts of the rat epididymis, only the 'long' mRNA molecules of the cauda region were efficiently translated and the antigen glycosylated, indicating that expression of rat CD52 is regulated on a post-transcriptional level. Lectin binding and deglycosylation studies supported the contention that there is extensive mucin-type O-glycosylation of rat CD52. In rats, there was no indication of complex N-linked carbohydrates similar to those described for human CD52.
IHF Institute for Hormone and Fertility Research at the University of Hamburg, Germany.
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Derr
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Yeung
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Cooper
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Kirchhoff
D000818:Animals; D015703:Antigens, CD / Q000032:analysis / Q000235:genetics* / Q000378:metabolism*; D000951:Antigens, Neoplasm; D015152:Blotting, Northern; D015153:Blotting, Western; D000074301:CD52 Antigen; D004822:Epididymis / Q000166:cytology*; D005434:Flow Cytometry; D019084:Fluorescent Antibody Technique, Indirect; D005786:Gene Expression Regulation; D006023:Glycoproteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism*; D006031:Glycosylation; D037102:Lectins / Q000378:metabolism; D008297:Male; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054801:Phosphoinositide Phospholipase C; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D013094:Spermatozoa / Q000276:immunology*; D010738:Type C Phospholipases / Q000378:metabolism
D015703:Antigens, CD; D000951:Antigens, Neoplasm; D000074301:CD52 Antigen; C000613862:CD52 protein, human; C000613864:Cd52 protein, rat; D006023:Glycoproteins; D037102:Lectins; D012333:RNA, Messenger; D010738:Type C Phospholipases; D054801:Phosphoinositide Phospholipase C; D043265:Phosphatidylinositol Diacylglycerol-Lyase