RING OR ZINC FINGER PROTEINS DURING GAMETOGENESIS AND EMBRYONIC DEVELOPMENT

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。生殖发育和功能是复杂的过程，既涉及基因决定的事件，也涉及生理事件。识别参与调控这些过程的关键基因是必要的，以确定这些过程是如何协调的。特别是，成熟配子的产生涉及许多发育和分化步骤，以及独特的细胞减数分裂过程，统称为配子发生。一般而言，分化和发育以及导致细胞表型获得和维持的其他过程依赖于基因表达的差异和协调调节。然而，精原干细胞的增殖、减数分裂停止和退出的信号、精子发生以及生殖细胞与相邻体细胞之间的相互作用等配子发生过程中基因表达的调控尚不清楚。此外，母系和父系基因在合子发育中的作用也没有很好的界定。锌指蛋白等生殖细胞特异性分子在配子发生和早期胚胎发育过程中的鉴定和功能鉴定将促进我们对配子和合子发育的了解，并将为开发生殖疾病诊断试剂、治疗不育症、性腺癌、出生前死亡和新型避孕药提供新的线索。环(真正有趣的新基因)指是一种特殊类型的锌指蛋白。它包含一个进化保守的结构，在人类基因组数据库的300多个蛋白质中发现，在这个结构中，两个氨基酸环在碱基上被八个半胱氨酸或组氨酸残基连接在一起，这些残基结合了两个锌离子。有两种不同的变种，C3HC4型和C3H2C3型，尽管半胱氨酸/组氨酸模式不同，但这两种类型显然是相关的。这些环指结构域主要存在于两类蛋白质中：(1)转录激活因子、抑制因子或辅助因子，以及(2)调节染色质的复合体的亚单位。这些蛋白质可能与其他蛋白质相互作用，参与泛素化，并在细胞生长控制中发挥作用，如细胞凋亡、肿瘤发生、DNA损伤修复和基因表达。计算机分析表明，大量与人类同源的环/锌指蛋白在生殖细胞或早期胚胎中表达，表明这些蛋白可能在生殖细胞和合子发育中发挥作用。最近的研究表明，环指蛋白、受精卵停滞1(ZAR1)和NEURL分别对受精卵发育、男性生育和怀孕期间的乳腺成熟起关键作用。去除另一种环指蛋白Siah1a也会导致男性不育，原因是减数分裂存在缺陷。此外，另一种卵母细胞特有的环指蛋白RFPL4与卵母细胞蛋白相互作用，可能作为E3泛素蛋白连接酶来调节蛋白质降解和减数分裂细胞周期进程。因此，我们才刚刚开始了解这些环/锌指蛋白在配子发生和早期胚胎发育中的功能作用。在搜索了小鼠和人类基因组数据库后，我发现七个未鉴定的基因可能在配子发生和合子发育过程中发挥作用，因为这些基因的表达序列标签(ESTs)存在于小鼠、人类生殖细胞或早期胚胎的cDNA文库中。我对这些基因中的五个(命名为未知1-5)特别感兴趣，它们位于不同的小鼠染色体上，都有人类的同源基因。UNKNOWN-1、UNKNOWN-2和UNKNOWN-3是C3HC4环指蛋白，UNKNOWN？4和UNKNOWN？5是C2H2锌指蛋白，类似于雄性或雌性生殖细胞发育所需的zfp148、Nanos2和nanos3。我将结合遗传学、分子和细胞方法来解决这些基因是否参与配子发生和早期胚胎发育。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Reproductive development and function are complex processes involving both genetically determined and physiological events. Identification of the critical genes involved in regulating these processes is necessary to characterize how these processes are coordinated. In particular, the generation of mature gametes involves many steps of development and differentiation, as well as the distinctive cellular process meiosis, that are collectively termed gametogenesis. In general, differentiation and development, and other processes that lead to the acquisition and maintenance of a cellular phenotype are dependent on the differential and coordinate regulation of gene expression. However, the regulation of gene expressioin many aspects of gametogenesis such as proliferation of spermatogonial stem cells, signals for meiotic arrest and meiotic exit, and spermiogenesis, and cross-talk between germ cells and adjacent somatic cells, are not clearly understood. In addition, the roles of maternal and paternal genes in zygotic development are also not well defined. Identification and functional characterization of germ cell-specific molecules such as zinc finger proteins during gametogenesis and early embryonic development would advance our knowledge regarding gamete and zygotic development, and will provide new leads for development of diagnostic reagents for reproductive diseases, novel therapeutic medicines for the treatment of infertility, gonadal cancers, pre-natal death, and novel contraceptive agents. The RING (Really interesting novel genes) finger is a specialized type of zinc finger proteins. It contains an evolutionarily conserved structure found in more than 300 proteins from the human genome database, in which two loops of amino acids are pulled together at their base by eight cysteine or histidine residues that bind two zinc ions. There are two different variants, the C3HC4-type and the C3H2C3-type, which is clearly related despite the different cysteine/histidine patterns. These ring finger domains are found in two major classes of proteins: (1) transcriptional activators, repressors or cofactors and (2) subunits of complexes that modulate chromatin. These proteins likely interact with other proteins, participate in ubiquitination, and play roles in cell growth control such as apoptosis, tumorigenesis, DNA damage repair, and gene expression. In silico analyses have demonstrated that a large number of ring/zinc finger proteins with human homologs are expressed in the germ cells or early embryos, indicating that these proteins may play a role in germ cell and zygotic development. Recent studies have shown that ring finger proteins, zygote arrest 1 (Zar1) and NEURL are critical for zygotic development, and male fertility and mammary gland maturation during pregnancy, respectively. Ablation of another ring finger protein, Siah1a, also caused male infertility due to defects in meiosis. In addition, another oocyte specific ring finger protein, RFPL4, interacts with oocyte proteins and likely functions as an E3 ubiquitin protein ligase to regulate protein degradation and meiotic cell cycle progression. Therefore, we are just in the beginning to understand the functional roles of these ring/zinc finger proteins during gametogenesis and early embryonic development. Having searched mouse and human genomic databases, I found that seven uncharacterized genes likely play a role during gametogenesis and zygotic development, as the expressed sequence tags (ESTs) of these genes are present in the cDNA libraries of mouse, human germ cells or early embryos. I am particularly interested in five of these genes (named unknown 1-5), which are located in different mouse chromosomes and all have human orthologs. Unknown-1, -2 and -3 are C3HC4 ring finger proteins, while unknown?4 and ?5 are C2H2 zinc finger proteins, similar to zfp148, nanos2 and nanos3, which are required for male or female germ cell development. I will combine genetic, molecular, and cellular approaches to address whether these genes are involved in the gametogenesis and early embryonic development.