Oct-4 Regulates Embryonal Carcinoma Differentiation

描述(申请人提供)：转移性胚胎癌是生殖细胞肿瘤(GCT)的一种有趣的特征，它能够分化为局限性畸胎瘤，后者由更成熟、生长较慢的外胚层、中胚层和内胚层三种胚胎谱系的组织组成。胚胎性癌(EC)的分化受调控基因的控制，这些基因介导永久性的表型改变。这些调控基因通常是转录调节器，激活或抑制基因表达模式，从而造成干细胞分化过程中出现的表型变化。这些转录因子不仅可以在GCT的特定分化阶段介导表型成熟，而且还可以调节在下一阶段分化中重要的转录因子的表达。这项资助的目的是增加对EC细胞中存在的转录因子如何调节EC细胞在第一次分化时做出的初始谱系决定的了解。POU同源结构域蛋白Oct-4和我们分离的Forkhead Box蛋白FoxD3(以前的Genesis)是在EC细胞中优先表达的转录因子。在正常的胚胎相当于EC细胞的胚胎干细胞(ES细胞)中，OCT-4在原肠形成过程中下调对于正常的谱系发育是必不可少的。我们之前已经发现Oct-4还可以作为辅助抑制因子，阻止FoxD3激活分化转录因子FoxA1和FoxD2的启动子。本研究将从三个特定的目的探讨Oct-4和FoxD3在EC细胞分化决策中的作用。1)对Oct-4与FoxD3的相互作用结构域进行定位，并对其功能进行实时蛋白质组学分析。2)研究Oct-4抑制谱系特异性转录激活的生化机制。3)研究Oct-4的抑制活性如何介导EC细胞的全潜能和畸胎瘤分化。

DESCRIPTION (provided by applicant): An intriguing characteristic of metastatic embryonal carcinoma, a type of germ cell tumor (GCT), is that they are able to differentiate to localized teratoma, which is made up of more mature, slower growing tissue of all three embryonic lineages, ectoderm, mesoderm, and endoderm. Differentiation of embryonal carcinoma (EC) is controlled by regulatory genes that mediate permanent phenotypic change. These regulatory genes are often transcriptional regulators that activate or repress patterns of gene expression that create the phenotypic change seen during stem cell differentiation. These transcription factors can not only mediate phenotypic maturation during a particular differentiation stage of GCT, but can also regulate expression of the transcription factors that are important in the next stage of differentiation. The goal of this grant is to increase understanding how the transcription factors present in the EC cell regulate the initial lineage decisions an EC cells makes as it first differentiates. The POU homeodomain protein Oct-4 and the Forkhead Box protein we isolated, FoxD3 (previously Genesis), are transcription factors preferentially expressed in EC cells. In the normal embryonic equivalent of EC cells, embryonic stem (ES) cells, downregulation of Oct-4 during gastrulation is essential for proper lineage development. We have previously found that Oct-4 can also act as a co-repressor to prevent FoxD3 from activating the promoters of the differentiation transcription factors FoxA1 and 2. This study will investigate the role of Oct-4 and FoxD3 in EC cell differentiation decisions in three specific aims. 1) The interacting domains between Oct-4 and FoxD3 will be mapped and their function analyzed in real time proteomics. 2) The biochemical mechanism by which Oct-4 represses lineage-specific transcriptional activation will be investigated. 3) How the repressor activity of Oct-4 mediates totipotentiality versus teratoma differentiation in EC cells will be studied.