Molecular Basis of BMP-15 Function

BMP-15 功能的分子基础

• 批准号: 7038282
• 负责人: SHUNICHI SHIMASAKI
• 金额: $ 33.4万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-05-01 至 2008-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7038282
• 关键词: biological signal transduction; bone morphogenetic proteins; follicle stimulating hormone; gene expression; gene mutation; genetically modified animals; granulosa cell; growth factor receptors; hormone receptor; laboratory mouse; laboratory rat; oogenesis; peptide chemical synthesis; protein biosynthesis; protein purification; protein structure function; receptor binding; receptor expression; site directed mutagenesis; southern blotting; transfection /expression vector; transforming growth factors

DESCRIPTION (provided by applicant): Bone Morphogenetic Protein-15 (BMP-15), a member of the TGF-beta superfamily, is an oocyte-specific growth factor. Recent studies of a naturally occurring mutation in sheep [Inverdale (FecX')] have suggested an essential role for BMP-15 in folliculogenesis and fertility. In the homozygous FecX' female, follicular development arrests at the primary stage, resulting in infertility. In contrast, the heterozygous FecX1 females exhibit increases in ovulation rate and in twin and triplet births. It has now been shown that a substitution of valine by aspartic acid at residue 31 (V31D) of the BMP-15 gene is present in FecX' carriers. Therefore, BMP-15 mutation appears to be associated with infertility and super fertility in a dosage-sensitive manner. Our laboratory has recently produced recombinant BMP-15 and identified granulosa cells (GCs) as target cells for this factor. Using the recombinant protein, we have found that BMP-15 (i) stimulates rat GC proliferation, (ii) inhibits FSH receptor expression, and (iii) stimulates kit ligand expression in the GCs. These findings establish the new concept that oocyte-derived BMP-15 is functionally involved in regulating three major aspects of follicle development, namely GC proliferation, GC cytodifferentiation, and oogenesis. Here, we propose to elucidate the molecular basis of BMP-15 action from the receptors to the target genes by characterizing BMP-15 receptor and signaling molecules (Aim 1) and the molecular mechanisms by which BMP-15 regulates FSH receptor and kit ligand gene expression (Aim 2). We have also found that, despite lacking the inter-subunit disulfide bonds seen in most of the dimeric TGF-beta superfamily members, BMP-15 and the closely related GDF-9 are homodimers. Furthermore, we have obtained evidence that BMP-15 and GDF-9 can form heterodimers, and thus we propose to explore the biological roles of BMP-15/GDF-9 heterodimers (Aim 3). Although the V31D mutation in the BMP15 gene was found in Inverdale sheep, the role of BMP-15-V31D mutation in Inverdale sheep is yet unclear. We will explore the biological activities of the BMP-15-V31D mutation and test our hypothesis that BMP-15-V31D could influence the biosynthesis of GDF-9 (Aim 4). Finally we will generate the same mutation in mice and determine the in vivo effect of the BMP-15-V31D mutation on folliculogenesis and ovulation (Aim 5). These studies should provide significant advances in understanding regulation of the events central to female fertility and ultimately improve approaches for birth control and for treating gonadal abnormalities and infertility.

描述（由申请人提供）：骨形态发生蛋白15（BMP-15），A TGF-β超家族的成员是卵母细胞特异性生长因子。最近的 研究绵羊天然发生的突变[inverdale（fecx'）] 提出了BMP-15在卵泡发生和生育方面的重要作用。在 纯合子的女性，卵泡发育在初级 阶段，导致不育。相反，杂合的FECX1女性 排卵率以及双胞胎和三胞胎出生的增加。现在有 已显示在残基31（v31d）的天冬氨酸取代valine BMP-15基因中存在于FECX的载体中。因此，BMP-15突变 似乎与不育和超级生育有关 剂量敏感的方式。我们的实验室最近产生了重组 BMP-15并将颗粒细胞（GC）鉴定为该因素的靶细胞。 使用重组蛋白，我们发现BMP-15（i）刺激大鼠GC 增殖，（ii）抑制FSH受体表达，（iii）刺激试剂盒 GCS中的配体表达。这些发现建立了一个新概念 卵母细胞衍生的BMP-15在功能上与调节三个主要 卵泡发育的各个方面，即GC增殖，GC 细胞分化和卵子发生。在这里，我们建议阐明分子 通过表征从受体到靶基因的BMP-15作用的基础 BMP-15受体和信号分子（AIM 1）和分子机制通过 BMP-15调节FSH受体和试剂盒配体基因表达（AIM 2）。我们 还发现，尽管缺乏在 大多数二聚体TGF-BETA超家族成员BMP-15和紧密 相关的GDF-9是同型二聚体。此外，我们获得了证据表明 BMP-15和GDF-9可以形成异二聚体，因此我们建议探索 BMP-15/GDF-9异二聚体的生物学作用（AIM 3）。虽然V31d 在Inverdale绵羊中发现了BMP15基因中的突变， Inverdale绵羊中的BMP-15-V31D突变尚不清楚。我们将探索 BMP-15-V31D突变的生物学活性，并检验我们的假设 BMP-15-V31D可能影响GDF-9的生物合成（AIM 4）。最后我们会的 在小鼠中产生相同的突变，并确定体内效应 BMP-15-V31D关于卵泡发生和排卵的突变（AIM 5）。这些研究 应该在理解事件的调节方面带来重大进展 女性生育的中心，最终改善了节育的方法 以及治疗性腺异常和不育。