Cloning of rat sp56, the homologue of mouse sperm ZP3 receptor-sp56.
Mouse sp56 is considered as one of the candidates for mouse zona pellucida 3 (mZP3) receptor. Up to date, its homologue has only been cloned from guinea pig, namely AM67. Based on the cDNA sequence of mouse sp56, we designed a pair of primer to amplify its homologue from rat testis cDNA. Using RT-PCR, two fragments of 743 bp and 938 bp were amplified. The PCR products show very high homology to mouse sp56. However, the 743 bp product completely lacks one of the seven Sushi domains of mouse sp56. Using the 743 bp product as the probe to detect the expression profile of sp56 in rat tissues, Northern blot shows that a approximately 2.0 kb mRNA expresses specifically in testis. Employed the RACE method, two full cDNA sequences of rat sp56 were obtained. A Mr approximately 42 KD band was detected in denatured and non-reducing protein sample of rat testis and sperm with anti-mouse sp56 monoclonal antibody by Western blot method. Rat sp56 was localized on rat sperm head by the indirect immunofluorescence method. Rat sp56 immunoreactivity was detected from the early pachytene spermatocytes and throughout the spermatogenesis. Its cloning will further our understanding of the mechanism of the sperm-egg recognition and binding.
Laboratory of Molecular Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China.
Xiao Bing
XB
He
Yuan Chang
YC
Yan
Yi Ping
YP
Li
S S
SS
Koide
D000818:Animals; D000906:Antibodies; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000032:analysis / Q000235:genetics; D004527:Egg Proteins / Q000378:metabolism*; D005306:Fertilization / Q000502:physiology*; D005455:Fluorescent Antibody Technique; D008297:Male; D008562:Membrane Glycoproteins / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D051381:Rats; D017208:Rats, Wistar; D011956:Receptors, Cell Surface / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D013094:Spermatozoa / Q000166:cytology / Q000378:metabolism*; D013737:Testis / Q000166:cytology / Q000378:metabolism*; D000072216:Zona Pellucida Glycoproteins
D000906:Antibodies; D018076:DNA, Complementary; D004527:Egg Proteins; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; D000072216:Zona Pellucida Glycoproteins; C000606521:Zp3 protein, mouse; C094480:sp56 protein, mouse