E2F AND APOPTOSIS

E2F 和细胞凋亡

• 批准号: 6706278
• 负责人: TIMOTHY F KOWALIK
• 金额: $ 28.7万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-02-05 至 2006-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6706278
• 关键词: apoptosis; brca gene; cell proliferation; cyclin dependent kinase; enzyme activity; gene induction /repression; gene mutation; nucleic acid sequence; oligonucleotides; oncoprotein p21; p53 gene /protein; tissue /cell culture; transcription factor

The long-term objectives of the proposed research are to define the molecular mechanisms underlying the normal relationship between proliferation and apoptosis and to understand the consequences of uncoupling this connection in diseases like cancer. Mounting evidence points to central roles of the Rb/E2F pathway in the commitment of cells to S phase and for p53 to limit the proliferative capacity of this pathway by inducing growth arrest or apoptosis. In support of this relationship, we have previously shown that expression of E2F1 not only leads to S phase induction, but also to a p53-dependent apoptosis. This apoptosis is specific to E2F1 and coincides with an E2F1-mediated accumulation of p53 protein. Both of these processes appear to be stimulated by E2F1 transcriptional activity. Using recombinant adenoviruses to efficiently express cDNAs in mouse embryo fibroblasts (MEFs) that are nullizygous for p19ARF, p53 or double null for Mdm2/p53, we now have evidence to suggest that E2F1 signals apoptosis and p53 accumulation through separate pathways. It has been suggested that E2F1 induces p53 accumulation and apoptosis by activating the p19ARF pathway. However, we find that although p19ARF is downstream of E2F1 in signaling p53 accumulation, p19ARF is not necessary for E2F1- mediated apoptosis. Interestingly, we do find that Mdm2 is required for both E2F1-mediated p53 accumulation and apoptosis. This implies that the p53 accumulation and apoptosis pathways diverge at the point where Mdm2 affects p53 function. To further investigate the different pathways by which E2F1 induces p53 accumulation and apoptosis, I plan to 1) characterize the pathway by which E2F expression leads to p53 protein accumulation and analyze the consequences of inducing this pathway; 2) determine the steps involved in a p19ARF-independent pathway that leads to p53-dependent apoptosis; and 3) identify apoptosis genes that are specifically induced by E2F1.

拟议的研究的长期目标是定义增殖与凋亡之间正常关系的分子机制，并了解在癌症等疾病中将这种联系解偶联的后果。 安装证据表明，RB/E2F途径在细胞对S相的承诺中的核心作用以及p53通过诱导生长停滞或凋亡来限制该途径的增殖能力。 为了支持这种关系，我们先前已经表明，E2F1的表达不仅会导致S相诱导，还导致p53依赖性细胞凋亡。 该细胞凋亡是E2F1的特异性，与E2F1介导的p53蛋白的积累相吻合。 这两个过程似乎都受到E2F1转录活性的刺激。 使用重组腺病毒在小鼠胚胎成纤维细胞（MEF）中有效表达cDNA，对于P19ARF，p53或Double null而言，对于MDM2/p53而言，我们现在有证据表明，E2F1信号的p53信号和p53通过单独的路径积累。 已经提出，E2F1通过激活P19ARF途径诱导p53的积累和凋亡。但是，我们发现，尽管p19arf在信号p53积累中是E2F1的下游，但对于E2F1介导的细胞凋亡并不需要P19ARF。 有趣的是，我们确实发现E2F1介导的p53积累和凋亡都需要MDM2。这意味着p53的积累和凋亡途径在MDM2影响p53功能的位置发出。 为了进一步研究E2F1诱导p53积累和凋亡的不同途径，我计划1）表征E2F表达导致p53蛋白积累并分析诱导该途径的后果的途径； 2）确定p19arf独立途径所涉及的步骤，该途径导致p53依赖性凋亡； 3）鉴定E2F1特异性诱导的凋亡基因。