Novel Pathway Involving E2F1, ATM and NBS1

描述（由申请人提供）：视网膜母细胞瘤（Rb）肿瘤抑制功能的丧失导致细胞增殖失控，并有助于大多数人类癌症的发展。在Rb失活的反应中，p53肿瘤抑制因子被激活，这通常导致细胞凋亡的消除。p53的这种“Rb守护者”功能在抑制肿瘤发生中起着关键作用。实验证据表明，Rb失活到p53活化的信号通路需要E2F1转录因子。E2F1的过表达或Rb失活导致p53积累并诱导凋亡。人们普遍推测，替代阅读框（ARF）肿瘤抑制因子是mdm2的一种抑制剂，可通过E2F1介导p53的激活。这一建议是基于E2F1转录激活ARF基因启动子的发现，以及其他癌基因，如Myc，需要ARF诱导p53依赖性细胞凋亡。在初步数据中，我们证明ARF对于e2f1诱导的细胞凋亡实际上是不可缺少的。相反，E2F1诱导p53依赖性细胞凋亡与咖啡因敏感的p53磷酸化有关。此外，失调毛细血管扩张突变激酶（ataxia-毛细血管扩张突变激酶）与DNA损伤激活p53有关，被发现是e2f1诱导的p53磷酸化和细胞凋亡所必需的。相反，e2f1刺激靶基因表达和促进S期进入的能力不受ATM缺失的影响。奈亨断裂综合征基因NBS1的产物也被证明是E2F1诱导p53磷酸化和细胞凋亡所必需的。NBS1是Mre11/Rad50 DNA修复复合体的一部分，最近被证明直接结合E2F1。这些发现扩展了ATM和NBS1的已知功能，并显著改变了当前关于细胞周期失调如何激活p53的模型。我们的假设是，不受管制的E2F1活性以nbs1依赖性的方式刺激ATM，从而激活p53和其他检查点反应因子。E2F1-ATM通路可能同时响应细胞周期失调和DNA损伤，从而诱导细胞凋亡和抑制肿瘤发生。这些研究的一个主要目标是从分子上确定ATM和NBS1在E2F1和p53之间的信号通路中的作用。p73和c-Abl在e2f1诱导的atm依赖性细胞凋亡中的作用也将被探讨。最后，我们将通过小鼠模型研究E2F1、ATM和NBS1在调节肿瘤发生中的功能关系。

DESCRIPTION (provided by applicant): Loss of retinoblastoma (Rb) tumor suppressor function leads to deregulated cell proliferation and contributes to the development of most human cancers. In response to Rb inactivation, the p53 tumor suppressor is activated and this usually results in the elimination of the cell by apoptosis. This "guardian of Rb" function for p53 plays a critical role in suppressing tumorigenesis. Experimental evidence demonstrates that the signaling pathway from Rb inactivation to p53 activation requires the E2F1 transcription factor. Deregulation of E2F1, by overexpression or Rb inactivation, results in p53 accumulation and the induction of apoptosis. It has been widely speculated that the alternative reading frame (ARF) tumor suppressor, an inhibitor of mdm2, mediates the activation of p53 by E2F1. This suggestion is based on the findings that E2F1 transcriptionally activates the ARF gene promoter and that other oncogenes, such as Myc, require ARF to induce p53-dependent apoptosis. In preliminary data we demonstrate that ARF is in fact dispensable for E2F1-induced apoptosis. Instead, the induction of p53-dependent apoptosis by E2F1 is correlated with the caffeine-sensitive phosphorylation of p53. Moreover, the ataxia-telangiectasia mutated (ATM) kinase, which has been implicated in activating p53 in response to DNA damage, is found to be required for E2F1-induced p53 phosphorylation and apoptosis. In contrast, the ability of E2F 1 to stimulate the expression of target genes and to promote S phase entry is unaffected by the absence of ATM. The product of the Nijmegen breakage syndrome gene, NBS1, is also shown to be required for E2F1 to induce the phosphorylation of p53 and apoptosis. NBS1 is part of the Mre11/Rad50 DNA repair complex and has recently been shown to directly bind E2F1. These findings expand the known functions for ATM and NBS1 and significantly alter the current model for how cell cycle deregulation activates p53. Our hypothesis is that deregulated E2F1 activity stimulates ATM in an NBS1-dependent manner to activate p53 and perhaps other checkpoint response factors. This E2F1-ATM pathway may respond to both cell cycle deregulation and DNA damage to induce apoptosis and suppress tumorigenesis. A major goal of these studies will be to molecularly define the roles of ATM and NBS1 in the signaling pathway between E2F1 and p53. A role for p73 and c-Abl in E2F1-induced, ATM-dependent apoptosis will also be explored. Finally, the functional relationship between E2F1, ATM and NBS1 in modulating tumor development will be examined using murine models.