Stage-Specific Germ Cell-Sertoli Cell Interactions

描述（由申请人提供）：发育中的雄性配子和邻近的支持细胞之间的相互作用对精子发生至关重要。支持细胞表达编码男性生殖细胞所需的细胞粘附分子、生长因子、转运蛋白、蛋白酶和蛋白酶抑制剂的基因。生殖细胞通过支持细胞调节许多这些基因的表达。因此，这些基因的表达随着邻近的胚芽在精原上皮周期的各个阶段的进展而改变。本应用程序从组织蛋白酶L基因的角度研究了支持细胞阶段特异性基因表达的调控和生物学后果。当邻近的生殖细胞从I期进展到VI期和VII期时，该基因的转录增加10倍以上，然后在生殖细胞达到x期时减少到无法检测到的水平。这种基因表达循环是对生殖细胞的反应，导致组织蛋白酶L基因转录的顺序抑制、刺激和再抑制。我们已经在组织蛋白酶L基因中发现了两个可能响应这些信号的结构域：[1]一个上游抑制结构域响应来自生殖细胞的抑制信号[2]一个120bp的激活结构域在睾丸成熟的高潮时被刺激，其特征是完成第一波精子发生。这些结构域在体内具有功能；组织蛋白酶L基因的3kb片段在转基因小鼠中同时具有支持细胞特异性和阶段特异性的报告基因表达。本研究还提出了组织蛋白酶L在精原上皮中的功能。这些实验是由于我们观察到表达催化失活组织蛋白酶L的小鼠表现出更高的精管萎缩发生率，并且在非萎缩小管中产生的精子比对照小鼠少30%。基于所有这些数据，本提案提出了四个问题，这是我们的具体目标：1激活域中特定的顺式作用元素是什么？它们的功能是什么？2. 与成熟结构域结合的转录因子的身份是什么？它们的表达是否有阶段特异性和成熟度依赖性？3. 支持细胞中哪些顺式作用元件和哪些生精细胞抑制组织蛋白酶L启动子活性？4. 组织蛋白酶L在精原上皮中的作用是什么？

DESCRIPTION (provided by applicant): Interactions between developing male gametes and adjacent Sertoli cells are essential for spermatogenesis. Sertoli cells express genes encoding cell adhesion molecules, growth factors, transport proteins, proteases and protease inhibitors required by male germ cells. Germ cells regulate expression by Sertoli cells of many of these genes. Consequently, expression of these genes changes as the adjacent germ progress through the stages of the cycle of the seminiferous epithelium. This application examines the regulation and biological consequences of stage-specific gene expression by Sertoli cells from the perspective of the cathepsin L gene. Transcription of this gene increases more than 10-fold as adjacent germ cells progress from stage I to stages VI and VII and then decreases to undetectable levels when germ cells reach stage X. This cycle of gene expression is a response to germ cells which causes sequential repression, stimulation and re-repression of transcription of the cathepsin L gene. We have identified 2 domains in the cathepsin L gene that potentially respond to these signals: [1] An upstream repressor domain responds to inhibitory signals from germ cells [2] A 120 bp activation domain is stimulated upon the culmination of testis maturation, which is characterized by the completion of the first wave of spermatogenesis. These domains are functional in vivo; a 3kb fragment of the cathepsin L gene confers both Sertoli cell-specific and stagespecific expression of a reporter in transgenic mice. This application also proposes to examine the function of cathepsin L in the seminiferous epithelium. Those experiments are prompted by our observation that mice which express catalytically inactive cathepsin L exhibit an increased incidence of seminiferous tubule atrophy and produce 30% fewer spermatids in nonatrophic tubules than are produced by control mice. Building on all of these data, this proposal asks four questions which are our specific aims: 1 What are the specific cis-acting elements in the activation domain and what are their functions? 2. What is the identity of the transcription factors that bind to the maturation domain? Is their expression stage-specific amd maturation-dependent? 3. What cis-acting elements and which spermatogenic cells repress cathepsin L promoter activity in Sertoli cells? 4. What is the function of cathepsin L in the seminiferous epithelium?