A 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，并导致大多数人类癌症的发生。作为对Rb失活的反应，P53肿瘤抑制基因被激活，这通常会导致细胞通过凋亡而被消除。P53的这种“Rb守护”功能在抑制肿瘤的发生中起着关键作用。实验证据表明，从Rb失活到P53激活的信号通路需要E2F1转录因子。通过过度表达或Rb失活来解除对E2F1的调控，会导致P53积聚并诱导细胞凋亡。人们普遍推测，MDM2的抑制因子ARF是一种肿瘤抑制因子，它介导了E2F1对P53的激活。这一建议是基于这样的发现，即E2F1转录激活ARF基因启动子，而其他癌基因，如Myc，需要ARF来诱导依赖于p53的细胞凋亡。在初步数据中，我们证明ARF实际上是E2F1诱导的细胞凋亡所必需的。相反，E2F1诱导的P53依赖的细胞凋亡与咖啡因敏感的P53的磷酸化有关。此外，共济失调-毛细血管扩张突变(ATM)激酶被发现是E2F1诱导的P53磷酸化和凋亡所必需的，它被认为是激活P53以应对DNA损伤的过程。相反，E2F1刺激靶基因表达和促进S进入时相的能力不受ATM的影响。Nijmegen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，最近被证明直接与E2F1结合。这些发现扩展了ATM和NBS1的已知功能，并显著改变了细胞周期放松如何激活P53的当前模型。我们的假设是，解除调控的E2F1活动以NBS1依赖的方式刺激ATM，激活P53，可能还有其他检查点反应因子。该E2F1-ATM通路可能同时对细胞周期紊乱和DNA损伤作出反应，从而诱导细胞凋亡和抑制肿瘤的发生。这些研究的一个主要目标将是从分子上确定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依赖的细胞凋亡中的作用也将被探索。最后，将使用小鼠模型研究E2F1、ATM和NBS1在调节肿瘤发展中的功能关系。

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.