MAMMALIAN GONADOGENESIS, FOLLICULOGENESIS AND FERTILIZATION

小鼠的受孕时间超过20天。在第7.25天的小鼠胚胎中首次检测到生殖细胞，在XX雌性小鼠中，生殖细胞从胚胎外的尿囊膜迁移到生殖器脊，在那里原始的性腺在第13天分化为卵巢。利用高通量测序和现代信息学方法筛选泌尿生殖系统cDNA文库，我们发现了一个位于X染色体上的新基因。该基因被命名为Nt2，在性别决定时优先在女性性腺的体细胞中表达。合成的蛋白质在其羧基末端附近含有一个同源盒域，这增加了NT2可能在性腺发生中发挥作用的可能性。该基因已成功定位于胚胎干细胞，目前正用于建立小鼠品系来分析无效表型。出生时，卵巢包含完整的生殖细胞，每个生殖细胞被一层颗粒细胞包围，这些颗粒细胞共同形成原始卵泡。我们已经鉴定出一种新的、卵母细胞特异的碱性螺旋-环-螺旋转录因子，FIG，即胚系中的因子。单拷贝Fig基因已经在胚胎干细胞中被破坏，这些细胞已经被用来建立小鼠品系。缺乏FIG的雌性小鼠是不育的，因为生殖细胞枯竭，继而无法在围产期形成原始卵泡。FIG下游靶点的识别将为卵泡形成的分子基础提供更多的见解。在卵泡形成开始后，FIG还调节编码ZP1、ZP2和ZP3的单拷贝基因的卵母细胞特异性表达(即在FIG缺失的小鼠中没有检测到带状转录本)。正常情况下，这三种透明带糖蛋白在卵泡发生过程中分泌形成透明带，这是一种细胞外基质，介导特定顺序的精子与卵子结合(例如，人类精子不会与小鼠透明带结合)。通过转基因，我们已经确定小鼠ZP1不是透明带形成、特定顺序精子结合或生育所必需的。相比之下，ZP2和ZP3都是形成基质所必需的，没有ZP3，小鼠就不能生育。尽管用人的同源物替换小鼠ZP2或小鼠ZP3可以恢复透明带基质，但这并不影响精子结合的特异性(即，小鼠的精子结合和受精卵被人化的透明带包围)。目前正在进行育种研究，以建立同时表达人类ZP2和ZP3或突变形式的小鼠ZP2和ZP3的小鼠品系，以进一步研究精子-卵子相互作用的分子基础。-透明带，受精，性别决定，卵母细胞特异性基因表达，生殖系中的因子，

Mouse gestation takes place over 20 days. Germ cells, first detected in mouse embryos at day 7.25, migrate from the extra-embryonic allantois to the genital ridge where the primitive gonad differentiates into an ovary by day 13 in XX females. Using high through-put sequencing and modern informatics to screen urogenital cDNA libraries, we have identified a novel gene located on the X chromosome. This gene, designated Nt2, is expressed preferentially in the somatic cells of female gonads at the time of sex determination. The resultant protein contains a homeobox domain near its carboxyl terminus raising the possibility that NT2 may play a role in gonadogenesis. The gene has been successfully targeted in embryonic stem cells that are currently being used to establish mouse lines for analysis of the null phenotype.At birth the ovary contains its full complement of germ cells, each surrounded by a single layer of granulosa cells which together form the primordial follicles. We have identified a novel, oocyte-specific, basic helix-loop-helix transcription factor, FIG (Factor In the Germline, alpha). The single-copy Fig gene has been disrupted in embryonic stem cells and these cells have been used to establish mouse lines. Female mice lacking FIG are sterile because of germ cell depletion secondary to an inability to form primordial follicles perinatally. Identification of downstream targets of FIG should provide additional insights into the molecular basis of follicle formation. After the onset of folliculogenesis, FIG also modulates the oocyte-specific expression of the single-copy genes that encode ZP1, ZP2 and ZP3 (i.e., zona transcripts are not detected in Fig null mice). Normally, the three zona glycoproteins are secreted during folliculogenesis to form the zona pellucida, an extracellular matrix that mediates order-specific sperm binding to the egg (e.g., human sperm will not bind to the mouse zona pellucida). Using transgenesis, we have determined that mouse ZP1 is not required for formation of the zona pellucida, order-specific sperm binding or fertility. In contrast, both ZP2 and ZP3 are required for matrix formation without which mice are infertile. Although the replacement of either mouse ZP2 or mouse ZP3 with their human homologue restores the zona pellucida matrix, it does not affect the specificity of sperm binding (i.e., mouse, but not human, sperm bind and fertilize eggs enclosed in humanized zonae pellucidae). Breeding studies to establish mouse lines expressing both human ZP2 and ZP3 or mutant forms of the mouse zona proteins are being pursued to further investigate the molecular basis of sperm-egg interactions. - zona pellucida, fertilization, sex determination, oocyte-specific gene expression, factor in the germline,