ANALYSIS FOR THE ROLE OF HUMAN SPERM ACROSOMAL MOLECULEH-SP-1 IN SIGNAL TRANSDUCTION VIA CALCIUM CHANNEL

在受精过程中，两个配子的几个表面分子参与配子识别(Sling，1989)。这些分子负责配子之间的受精和特定物种的相互作用的每个过程。对这些分子的研究对于了解精卵相互作用的机制和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cDNA文库中筛选出人精子抗原基因h-Sp-1。H-Sp-1基因序列分析表明，该基因全长2170个核苷酸，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…编码的分子更多的基因可能是一个跨膜蛋白，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。为了鉴定h-Sp-1基因编码的分子，用基于h-Sp-1基因的合成多肽(氨基酸编号174-198)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。我们还用电子显微镜分析了h-Sp-1分子的定位。结果表明，该分子存在于整个精子头表面。根据蛋白质印迹分析，该h-Sp-1似乎与钙调蛋白有关。活动。较少

Several surface molecules of both gametes are involved in gametes recognition during fertilization (Saling, 1989). These molecules are responsible for each process of fertilizationand 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-1) by screening from human testis cDNA library with human sperm specific antisera. The sequence analysis of the h-Sp-1 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 … More 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 characterizethe molecules encoded by the h-Sp-1 gene, the antisera were generated by immunizing rabbits with synthetic polypeptides (amino acid 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. We also analyzed the localization of h-Sp-1 molecule by electoromicroscpy. The result indicated the molecule exist on surface of whole sperm head. This h-Sp-1 seemed to be associaed with calmodulin, syntax in according to western blot analysis. activities. Less