Stage-Specific Germ Cell-Sertoli Cell Interactions

描述（由申请方提供）：发育中的雄性配子与相邻支持细胞之间的相互作用对于精子发生至关重要。支持细胞表达编码雄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。生殖细胞调节支持细胞中许多基因的表达。因此，这些基因的表达随着邻近的生殖细胞在生精上皮周期的各个阶段的进展而变化。本申请从组织蛋白酶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。当相邻的生殖细胞从I期进展到VI期和VII期时，该基因的转录增加超过10倍，然后当生殖细胞达到X期时降低到不可检测的水平。该基因表达循环是对生殖细胞的响应，其引起组织蛋白酶L基因转录的顺序抑制、刺激和再抑制。我们已经确定了组织蛋白酶L基因中的2个结构域，它们可能对这些信号产生响应：[1]上游阻遏物结构域对来自生殖细胞的抑制信号产生响应[2]一个120 bp的激活结构域在睾丸成熟达到顶点时受到刺激，其特征在于精子发生的第一波完成。这些结构域在体内是功能性的;组织蛋白酶L基因的3 kb片段赋予支持细胞特异性和阶段特异性的报告基因在转基因小鼠中的表达。本申请还提出检查组织蛋白酶L在生精上皮中的功能。我们观察到表达无催化活性的组织蛋白酶L的小鼠表现出生精小管萎缩的发生率增加，并且在非萎缩小管中产生的精子细胞比对照小鼠少30%。基于所有这些数据，这个建议提出了四个问题，这是我们的具体目标：1什么是特定的顺式作用元件在激活域和他们的功能是什么？2.与成熟结构域结合的转录因子的身份是什么？它们的表达是阶段特异性和成熟依赖性的吗？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在生精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?