X-linked Genes Involved In Early Mouse Embryonic Develop

X连锁基因参与早期小鼠胚胎发育

• 批准号: 6668116
• 负责人: DAVID SCHLESSINGER
• 金额: --
• 依托单位: NATIONAL INSTITUTE ON AGING
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6668116
• 关键词: apoptosis; developmental genetics; early embryonic stage; hypoparathyroidism; laboratory mouse; mammalian embryology; placenta; sex chromosomes

Systematic analyses of transcripts in early embryogenesis by the group of Dr. M. Ko in our Laboratory have uncovered a relative dearth of X-linked genes. Nevertheless, some X-linked genes show prominent effects in early development. Three of these have been chosen for special study. One turns out to be a member of the Bex family, Bex-3, that has not been previously isolated. The transcribed sequence is 789 bp in length and encodes a protein of 124 residues with a CAAX motif at its C-terminal end. It appears in a single cDNA and Northern species of 1.1 kb. Expression declines after the fetal period, but adult mouse and human show sustained expression in several organs, including brain, testis and ovary. The putative Bex-3 amino acid sequence shows very high homology to a partially characterized human cDNA clone called pHGR74, which is associated with spontaneous ovarian granulosa cell carcinoma. In vitro-expressed Bex3 is attached to the plasma membrane, and we are examining hints that it may be involved in signal transduction related to apoptosis during mouse development. A second gene, PLAC1, has important features that include: 1) expression with high specificity in placenta from early on in development; and 2) X-linkage very near the HPRT gene, at a location that makes it a candidate for involvement in placental dysplasia and placentomegaly when it is under- or overexpressed. It is one of very few genes that show very greatly increased activity in cloned animals, associated with placentomegaly. We have analyzed the structure of the cDNA and genomic form of PLAC1, and have shown that it is localized in giant cells and labyrinthine trophoblasts in the developing placenta, between 7.5 and 14.5 days post coitum. In additional studies with collaborators, we have found that the gene is comparably expressed in human placenta, but strong expression lasts longer, all the way to term. Currently we have started to analyze the basis for tissue-specific expression of the gene. Extensive transfections with constructs containing increasing amounts of DNA upstream of the transcription unit, fused to luciferase have thus far shown that neither 3.3 nor 7.5 kb of upstream DNA is sufficient to support the transcription of the gene in choriocarcinoma cells that express the endogenous PLAC1 gene. We interpret our results to mean that the specific expression of the gene involves more distant upstream (or intragenic!) elements, and are continuing to study increasingly large genomic regions to define regulatory elements more precisely. Additional novel genes that are uniquely expressed in placenta have been identified, and two of them have been studied to look for common regulatory features in their promoters and that of PLAC1.

我们实验室的M.Ko博士团队对早期胚胎发育的转录本进行了系统分析，发现相对缺乏X连锁基因。然而，一些X连锁基因在早期发育中表现出显著的作用。其中三个已被选作特别研究。其中一人被证明是Bex家族的成员，Bex-3，以前从未被隔离过。该转录序列全长789个碱基，编码124个残基的蛋白质，其C-末端带有CAAX基序。它存在于1.1kb的单一cDNAs和Northern种中。胎儿期后表达下降，但成年鼠和人在几个器官中持续表达，包括大脑、睾丸和卵巢。推测的Bex-3氨基酸序列与与自发性卵巢颗粒细胞癌相关的部分特征的人cDNA克隆PHGR74有很高的同源性。在体外表达的Bex3附着在质膜上，我们正在检查它可能参与小鼠发育过程中与细胞凋亡相关的信号转导的线索。第二个基因PLAC1具有重要的特征，包括：1)在胎盘发育的早期就有高度特异性的表达；以及2)与HPRT基因非常接近的X-连锁，当HPRT基因表达过低或过高时，它可能参与胎盘发育不良和胎盘肥大。它是极少数在克隆动物中表现出极大活性增加的基因之一，与胎盘肥大有关。我们对PLAC1的结构和基因组形式进行了分析，结果表明，PLAC1定位于胎盘发育中的巨细胞和迷路滋养细胞中，时间为7.5~14.5天。在与合作者的进一步研究中，我们发现该基因在人类胎盘中也有类似的表达，但强表达持续时间更长，一直持续到足月。目前，我们已经开始分析该基因组织特异性表达的基础。到目前为止，用含有越来越多的转录单位上游DNA的构建体与荧光素酶融合的广泛转染表明，3.3或7.5kb的上游DNA都不足以支持表达内源性PLAC1基因的绒毛膜癌细胞中该基因的转录。我们解释说，我们的结果意味着该基因的特定表达涉及更远的上游(或基因内！)正在继续研究越来越大的基因组区域，以更准确地定义调控元件。其他在胎盘中唯一表达的新基因已经被发现，其中两个已经被研究，以寻找它们的启动子和PLAC1的共同调节功能。