IDENTIFICATION/CHARACTERIZATION OF POTENTIAL NOVEL MATERNAL-EFFECT GENE PRODUCTS

这个子项目是许多利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 我们已经确定了一组六个NLRP（NLR家族，pyrin域包含）基因，在恒河猴卵母细胞和早期植入前胚胎中特异性表达。 小鼠NLRP 5已被证明是超过2-细胞阶段的胚胎发生所必需的，并且人NLRP 7被鉴定为第一个母体效应基因，因为其突变已与各种形式的生殖损耗相关联。 然而，其他NLRP家族成员的功能尚不清楚。 我们在非人灵长类动物中的初步数据，沿着最近在啮齿类动物和家畜中的发现，表明这些卵母细胞特异性NLRP基因在哺乳动物的卵子发生和早期胚胎发育过程中发挥重要作用。 本项目的目的是揭示小鼠和恒河猴卵母细胞特异性NLRPs的功能。 具体目标是：（1）使用敲除小鼠模型表征小鼠NLRP 4A和9 B的体内功能;（2）证明小鼠和猕猴卵母细胞中所选NLRP基因（小鼠NLRP 4a、5和9 b;猕猴NLRP 4、5和9）的体外排卵后作用;（3）鉴定和分析小鼠和猕猴卵母细胞中与所选NLRP蛋白相互作用的因子。 NLRP蛋白含有一个Pyrin结构域（PYD）、一个NACHT和一个富含亮氨酸的重复序列（LRR）结构域; PYD和LRR都介导蛋白质-蛋白质相互作用。 实验设计包括基因敲除小鼠模型的建立和分析、小鼠和猴卵母细胞中的RNA干扰（RNAi）、酵母双杂交系统在小鼠和猕猴卵母细胞中的应用。

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. We have identified a group of six NLRP (NLR family, pyrin domain containing) genes that are specifically expressed in the oocyte and early preimplantation embryos in rhesus macaques. Mouse NLRP5 has been demonstrated to be essential for embryogenesis beyond the 2-cell stage, and human NLRP7 was identified as the first maternal effect gene as the mutations of which have been linked to various forms of reproductive wastage. However, functions of other NLRP family members are not known. Our preliminary data in nonhuman primates, along with recent findings in rodents and domestic animals, suggest that these oocyte-specific NLRP genes play important roles during oogenesis and early embryo development in mammals. The purpose of this project is to reveal the functions of selected oocyte-specific NLRPs in mice and rhesus macaques. The specific aims are to: (1) characterize in vivo functions of mouse NLRP4A and 9B using knockout mouse models; (2) demonstrate the in vitro postovulatory roles of selected NLRP genes (mouse Nlrp4a, 5 and 9b; macaque NLRP4, 5 and 9) in mouse and macaque oocytes; (3) identify and analyze factors that interact with selected NLRP proteins in mouse and macaque oocytes. NLRP proteins contain a pyrin domain (PYD), a NACHT, and a leucine rich repeat (LRR) domain; both the PYD and LRR mediate protein-protein interactions. Experimental designs include generation and analysis of gene knockout mouse models, RNA interference (RNAi) in mouse and monkey oocytes, and application of yeast two-hybrid system in the mouse and macaque oocyte.