ZEB/deltaEF1 and Estrogen Signaling Cascades

说明(申请人提供)：除了其正常的生理功能外，雌激素还能引导导致保护和促进许多病理情况的事件。然而，将受体与其目标基因的结合与最终的生物学后果联系起来的分子细节往往还不清楚。为了确定雌激素触发的信号级联反应，有一个新的观察发现，ZeB/Def1基因是由雌激素在转录水平上被诱导的。另有数据表明，ZeB/Def1是生殖组织中雌激素信号转导途径中的一个环节。例如，Zeb/Def1在快速增殖的子宫内膜癌和卵巢癌中高表达，但不再受雌激素调节。相比之下，在生长缓慢的子宫内膜癌中，该基因被缺失。这些数据表明，ZeB/deltaEF1在生殖组织中起着关键作用。ZeB/deltaEF1是一种转录因子，含有多个功能结构域，每个末端有两个锌指簇。大多数数据表明，ZeB/deltaEF1是一个转录抑制因子，但在某些情况下，它作为转录激活因子发挥作用。该提案的总体目标是确定ZeB/deltaEF1激活基因的机制，并确定其靶基因。这项建议的具体目标是1)。探讨ZeB/deltaEF1，2)基因激活的机制。确定生殖组织中与ZEBdeltaEF1共同激活的蛋白质，以及3)。确定乳腺癌细胞系中ZeB/deltaEF1的下游靶点。第一个目标将主要通过截断ZeB/Def1并确定对报告基因激活的影响来实现。第二个具体目标将通过两种互补的方法来实现，串联质谱仪和酵母双杂交RAS招募系统。对于最后一个特定目的，将使用DNA微阵列的基因图谱来确定ZeB/deltaEF1的目标基因。这些实验将为研究ZeB/deltaEF1的基因激活机制和雌激素信号转导途径提供重要信息。

DESCRIPTION (provided by applicant): In addition to its normal physiological functions, estrogen directs events that lead to both the protection from and promotion of many pathological conditions. However, the molecular details linking the binding of the receptor to its target genes with the ultimate biological consequence often remain unclear. In an attempt to define signaling cascades triggered by estrogen, the novel observation was made that the ZEB/dEF1 gene is induced at the transcriptional level by estrogen. Other data show that ZEB/dEF1 is one of the links in estrogen signal transduction pathways in reproductive tissues. For example, ZEB/dEF1 expression is high in rapidly proliferating endometrial and ovarian cancers but is no longer regulated by estrogen. In contrast, the gene is deleted in slowly growing endometrial cancers. These data suggest that ZEB/deltaEF1 plays a pivotal role in reproductive tissues. ZEB/deltaEF1 is a transcription factor that contains several functional domains including two zinc finger clusters at each terminus. Most data suggest that ZEB/deltaEF1 is a transcriptional repressor but in some circumstances it functions as a transcriptional activator. The overall goals of this proposal are to define the mechanism of gene activation by ZEB/deltaEF1 and to identify its target genes. The specific aims of this proposal are to 1). Investigate the mechanism of gene activation by ZEB/deltaEF1, 2). Identify the proteins in reproductive tissues that act as co-activators with ZEBdeltaEF1, and 3). Identify the downstream targets of ZEB/deltaEF1 in a breast cancer cell line. The first aim will be approached primarily by truncating ZEB/dEF1 and determining the effects on reporter gene activation. The second specific aim will be addressed by two complementary approaches, tandem mass spectrometry and a yeast two-hybrid Ras recruitment system. For the last specific aim, target genes for ZEB/deltaEF1 will be identified using gene profiling with DNA microarrays. These experiments will provide important information about the mechanism of gene activation by ZEB/deltaEF1 and about estrogen signal transduction pathways.