BIOREGULATION OF SPERMATOGENESIS

我们发现支持细胞分泌蛋白S70具有共同的抗原性 Sertoli细胞分泌蛋白多肽成分的同源性 S45-S35异二聚体蛋白。我们做了意想不到的观察 针对这两种支持细胞分泌的多克隆抗血清 蛋白质与精子尾部外密纤维多肽的交叉反应 并且针对主要的多肽产生的多克隆抗血清 精子尾部外层致密纤维与蛋白S70及其受体的交叉反应 细分产品。这些发现暗示了一种可能的产物--前体 S70和S45-S35异二聚体蛋白与RAGE的关系 关于细胞起源和分子的重要问题 精子尾部外层致密纤维多肽的组织。 确定S70是否是S45-S35异二聚体的前体 蛋白质，我们将分析S70和S45-S35的结构和表达 异源二聚体蛋白基因与基因组同源区域的寻找 在生精细胞中。针对这两种病毒产生的多克隆抗体 Sertoli细胞分泌蛋白将用于筛选大鼠睾丸基因 Lambda gtll噬菌体表达载体中的表达文库。插页 从阳性克隆将被亚克隆到一个多功能质粒中 载体测定全长cdna序列并衍生蛋白质氨基酸 酸序列。推导出的氨基酸序列将与N- S70蛋白和外密蛋白的末端氨基酸序列分析 纤维多肽。CDNA将用于原位细胞杂交和 Northern印迹实验确定生精细胞， 主要是精子细胞，包含基因组序列和转录本 补充Sertoli细胞分泌蛋白。 确定支持细胞分泌蛋白是否与 外膜致密纤维多肽，我们将提高多克隆抗体 抗精子尾部纯化的外密纤维多肽和 寻找交叉反应表位和N末端氨基酸序列 Sertoli细胞分泌蛋白S70和S45-S35异二聚体的同源性 蛋白质和外层致密纤维多肽。亚细胞 支持细胞分泌蛋白和外膜蛋白的定位和去向 免疫胶体金法检测大鼠睾丸中致密纤维多肽 电子显微镜。最终，个体发育、荷尔蒙和 支持细胞分泌蛋白S70和S70对生精细胞的依赖性 S45-S35异源二聚体蛋白基因的活性和表达 大鼠睾丸发育过程中和培养的Sertoli的测定 使用自动细胞灌流系统的细胞 放射免疫分析。

We have found that Sertoli cell secretory protein S70 share antigenic homology with polypeptide components of Sertoli cell secretory protein S45-S35 heterodimeric protein. We have made the unexpected observations that polyclonal antisera raised against these two Sertoli cell secretory proteins crossreact with outer dense fiber polypeptides of sperm tail and that a polyclonal antiserum generated against a major polypeptide of outer dense fibers of sperm tail crossreacts with protein S70 and its breakdown products. These findings suggest a possible product-precursor relationship between S70 and S45-S35 heterodimeric protein and raise important questions concerning the cell origin and molecular organization of outer dense fiber polypeptides of the sperm tail. To determine whether S70 is a precursor for S45-S35 heterodimeric protein, we will analyze the structure and expression of S70 and S45-S35 heterodimeric protein genes and search for regions of genomic homology in spermatogenic cells. Polyclonal antibodies raised against these two Sertoli cell secretory proteins will be used to screen rat testis cDNA expression libraries in lambda gtll phage expression vector. Inserts from positive clones will be subcloned into a multifunctional plasmid vector to determine the complete cDNA sequence and derive protein amino acid sequence. Derived amino acid sequence will be compared with N- terminus amino acid sequence analysis of protein S70 and outer dense fiber polypeptides. cDNA will be used for in situ cytohybridization and Northern blot experiments to determine whether spermatogenic cells, mainly spermatids, contain genome sequences and transcripts complementary to Sertoli cell secretory proteins. To determine whether Sertoli cell secretory proteins are related to outer dense fiber polypeptides, we will raise polyclonal antibodies against purified outer dense fiber polypeptides from sperm tail and search for crossreactive epitopes and N-terminus amino acid sequence homology of Sertoli cell secretory protein S70 and S45-S35 heterodimeric protein and outer dense fiber polypeptides. The subcellular localization and fate of both Sertoli cell secretory proteins and outer dense fiber polypeptides will be determined in rat testis by immunogold electron microscopy. Ultimately, the ontogeny, hormonal and spermatogenic cell dependency of Sertoli cell secretory proteins S70 and S45-S35 heterodimeric protein gene activity and expression will be determined in rat testis during development and in cultured Sertoli cells using an automated cell perifusion system combined with radioimmunoassay.