Developing shRNA vectors to study the activating E2Fs

描述(申请人提供)：在大多数人类癌症中，肿瘤抑制基因Rb在功能上是失活的，随之而来的E2F家族成员及其转录靶点的解除调控被认为是肿瘤发生的关键步骤。根据其相反的转录特性，E2F可分为两类：激活型E2F和抑制性E2F。关于激活的和抑制的E2F在控制正常和肿瘤细胞增殖中的相对作用一直存在很大的争议。此外，这些蛋白质在ES细胞调节中的作用还没有被解决。这项提议的目标是创建一种新型的shRNA载体系统，该系统将能够在培养细胞或小鼠模型中诱导多个E2F被击倒。这些研究将使我们能够直接解决在ES细胞、原代成纤维细胞和肿瘤发展中激活E2F的需求。此外，它还将为社区提供一个有价值的载体资源，可用于高效、同时敲除所选择的基因。

DESCRIPTION (provided by applicant): The tumor suppressor Rb is functionally inactivated in most human, cancers and the consequent deregulation of E2F family members and their transcriptional targets is believed to be a crucial step in tumorigenesis. Based on their opposing transcriptional properties, the E2Fs can be divided into two classes: the activating E2Fs and the repressive E2Fs. There has been much debate about the relative roles of the activating versus repressive E2Fs in controlling the proliferation of both normal and tumor cells. Moreover, the role of these proteins in ES cell regulation has not yet been addressed. The goal of this proposal is to create a novel shRNA vector system that will enable inducible knockdown of multiple E2Fs in either cultured cells or mouse models. These studies will allow us to directly address the requirement of the activating E2Fs in ES cells, primary fibroblasts and in tumor development. Moreover, it will also provide a valuable vector resource to the community that can be used for efficient, simultaneous knockdown of genes of choice.