MOLECULAR ANALYSIS OF HUMAN SPERM ACROSOMAL MOLECULE H-SP-1 IN SPERM-EGG INTERACTION

在受精过程中，两个配子的几个表面分子参与配子识别(萨玲，)。这些分子负责受精的每个过程和配子之间特定物种的相互作用。对这些分子的研究对于了解精子与卵子相互作用的机制以及避孕疫苗的研制具有重要意义。本研究以人精子特异性抗血清为模板，从人睾丸c DNA文库中筛选得到人精子抗原基因h-Sp-L。序列分析表明，h-Sp-L基因全长2170bp，与人突触素基因有45%的同源性，被认为在神经末梢突触小泡的钙依赖信号转导中起重要作用。对h-Sp-1基因推导的氨基酸序列分析表明，该基因编码259个氨基酸，包括4个亲水区和4个疏水区。这表明h-Sp-1基因编码的分子可能是一种跨膜蛋白，与突触素一样具有四个跨膜区域。Northern杂交分析表明，h-Sp-1基因有2.3kb和1.1kb两种转录本。2.3kb的转录本在人睾丸中普遍表达，1.1kb的转录本在人睾丸中大量表达。为了研究h-Sp-L基因编码的分子特征，用人工合成的h-Sp-1基因多肽(氨基酸第174-198号)免疫兔制备抗血清。Western印迹分析表明，该抗血清与40 kDa的分子发生反应。该抗血清还对固定的人精子头进行了间接免疫荧光染色。纯化的抗血清对人精子穿透去透明带仓鼠卵子的实验有抑制作用。然而，它们没有表现出任何精子固定和凝集活性。这些结果提示h-Sp-1基因编码的分子可能在精子-卵子相互作用中起重要作用。

Several surface molecules of both gametes are involved in gametes recognition during fertilization( Saling, 1989 ). These molecules are responsible for each process of fertilization and species-specific interaction between the gametes. Studies of these molecules may be essential for understanding the mechanism of sperm-egg interaction and for the development of a contraceptive vaccine.We isolated a human sperm antigen gene (h-Sp-l) by screening from human testis cDNA library with human sperm specific antisera. The sequence analysis of the h-Sp-l gene indicated that it contained 2170 bp and had 45% homology to human synaptophysin gene, which was considered to play a crucial role in calcium dependent signal transduction in synaptic vesicles in the terminal end of nerves. The analysis of the deduced amino acids from the h-Sp- 1 gene showed that it encoded 259 amino acids containing four hydrophilic regions and four hydrophobic regions. This indicated that the molecules encoded by the h-Sp- 1 gene might be a transmembrane protein with four membrane spanning regions like the synaptophysin. Northern blot analysis indicated that h-Sp-1 gene had 2 kind of transcripts ( 2.3 kb and 1.1kb ). 2.3kb transcript expressed ubiquitously whereas 1.1kb transcript expressed abundantly in human testis. In order to characterize the molecules encoded by the h-Sp-l gene, the antisera were generated by immunizing rabbits with synthetic polypeptides ( aminoacid No 174-198 ) based on the h-Sp-1 gene. Western blot analysis indicated that the antisera reacted a 40 kDa molecule. The antisera also stained fixed human sperm head by indirect immunofluorescence staining. The purified antisera showed the inhibitory effect on human sperm penetration assay into zona-free hamster eggs. However, they did not show any sperm immobilizing and agglutinating activities. These results suggested that the molecules encoded by the h-Sp-1 gene might play an important role in the sperm-egg interaction.