Mouse models for pRB growth control via E2F/DP action

描述（由申请方提供）：人（RB）和小鼠（Rb）中视网膜母细胞瘤肿瘤抑制因子的失活促进肿瘤进展。在很大程度上，pRB通过抑制E2 F/DP家族复合物来调节生长，E2 F/DP家族复合物的一个子集在游离时刺激细胞周期进程。相反，pRB通过调节负责特定分化程序的转录因子（例如，MyoD、C/EBP、CBFA 1的激活）。然而，pRB的丢失并不损害所有组织的发育，并且生殖系RB或Rb突变导致高度渗透的、组织特异性的肿瘤倾向。在体外，G1细胞周期蛋白/Cdk介导的磷酸化的程度决定了pRB抑制细胞周期进程的能力;然而，最近证明G1细胞周期蛋白和Cdk对于大多数组织的正常发育是不可或缺的;提出了体内通常调节pRB/E2 F/DP途径的“布线”是什么的问题。可能的是，pRB的独特的肿瘤抑制功能（在pRB家族中）源于其协调生长抑制与特异性谱系定型的能力。只有通过识别在体内起作用的信号，我们才能理解Rb的缺失如何破坏关键组织的发育并促进特定部位的肿瘤发生。首先，使用我们新的一系列野生型和突变型Rb启动子转基因和新的Rb启动子敲入等位基因，我们将鉴定通常调节Rb的神经元特异性表达的激活子和阻遏子，并测试体外和体内Rb表达失调的功能后果（目的11.其次，我们将通过siRNA介导的DP家族表达的敲低和条件性Dpi和/或Dp 2敲除小鼠的建立，确定整个DP家族及其单个家族成员（Dpi和Dpi）在细胞周期控制和胚胎发育中的需求（目的2）。

DESCRIPTION (provided by applicant): Inactivation of the retinoblastoma tumor suppressor in humans (RB) and in mice (Rb) facilitates neoplastic progression. To a large part, pRB regulates growth by repressing E2F/DP famil) complexes, a subset of which when free, stimulates cell cycle progression. Instead, pRB stimulates differentiation by regulating transcription factors responsible for specific differentiation programs (e.g., activation of MyoD, C/EBP, CBFA1). Yet, loss of pRB does not compromise the development of all tissues, and germ-line RB or Rb mutations lead to highly penetrant, tissue specific tumor predisposition. In vitro, the extent of Gl Cyclin/Cdk-mediated phosphorylation governs the ability of pRB to restrain cell cycle progression; however, the recently demonstrated dispensability of Gl Cyclins and Cdks for the normal development of most tissues; raises the question of what is the "wiring" that normally regulates the pRB/E2F/DP pathway in vivo. It is likely that the unique tumor suppressive function of pRB (among pRB family numbers) stems from its ability to coordinate growth inhibition with specific lineage commitment. Only by identifying the signals operating in vivo can we understand how loss of Rb disrupts the development of key tissues and facilitates tumorigenesis at specific sites. First, using our novel series of wildtype and mutant Rb promoter transgenics and a new Rb promoter knock-in allele, we will identify the activators and repressors that normally regulate the neuronal-specific expression of Rb and test the functional consequences of deregulating Rb expression in vitro and vivo (Aim 11. Second, we will define a requirement for the entire DP family and its individual family members (Dpi and Dpi) in cell cycle control and embryonic development through siRNA-mediated knockdown of DP family expression and the establishment of conditional Dpi and/or Dp2 knockout mice (Aim 2).