ANALYSIS OF P73 FUNCTION AND REGULATION BY E2F 1

E2F 1对P73功能和调控的分析

• 批准号: 6377273
• 负责人: MEREDITH S IRWIN
• 金额: $ 10.25万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-08-01 至 2002-04-01
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6377273
• 关键词: apoptosis; cis platinum compound; doxorubicin; etoposide; flow cytometry; gene induction /repression; genetic transcription; immunofluorescence technique; messenger RNA; northern blottings; posttranslational modifications; protein structure function; tissue /cell culture; transcription factor; tumor suppressor proteins; western blottings

Dual inactivation of the retinoblastoma (pRb) and p53 tumor suppressor proteins occurs commonly during carcinogenesis. Inactivation of pRb leads to dysregulation of the E2F transcription factor family. E2F promotes both cell-cycle progression and apoptosis. The latter involves both p53- dependent and p53-independent mechanisms. My preliminary data suggests that E2F-1 can induce a recently identified p53 homolog called p73. p73 can, at least when overproduced, activate the transcription of p53-responsive genes and inhibit cell growth by inducing apoptosis. Since endogenous levels of p73 are very low and unlike p53, upstream signaling pathways leading to p73 activation have yet to be elucidated, it is intriguing that E2F-1 induces a significant increase in p73 levels. Dysregulation of the E2F/pRb pathway is common to most tumor cells. Therefore understanding the function of p73 within this pathway may provide clues as to the role of p73 in tumorigenesis. The goal of Specific Aim 1 is to determine the mechanism whereby E2F-1 induces p73. Standard protein stability and quantitative RNA assays will be employed to determine whether the effect of E2F-1 on p73 is due to changes in transcription, mRNA stability, and/or post-translational modifications. Whether the induction of p73 by E2F-1 in p53 nullizygous cells translates into activation of p53 target genes and apoptosis will form the basis of Specific Aims 2 and 3. The ability of E2F-1 to induce p73 dependent transactivation of p53 and p73 target genes will be assayed both biochemically and in transcription-based functional assays. In Specific Aim 3, flow cytometry and immunofluorescence techniques will be used to investigate whether p73 contributes to E2F-1 dependent apoptosis. p73 dominant negative proteins will be used to determine specificity in these assays. Finally, in Specific Aim 4 the role of p73 in E2F- dependent apoptosis induced by chemotherapeutic agents will be investigated. These studies may provide the first clues as to the signals that regulate p73 and may also help to explain p53 independent killing by E2F. Ultimately, the results of the studies proposed in this application may help in the development of novel gene therapies in which the E2F-1 protein, or perhaps, p73, can be used to induce tumor cell apoptosis.

视网膜母细胞瘤（pRb）和p53肿瘤抑制蛋白的双重失活通常发生在癌变过程中。pRb的失活导致E2F转录因子家族的失调。E2F促进细胞周期进程和细胞凋亡。后者涉及p53依赖性和p53非依赖性机制。我的初步数据表明，E2F-1可以诱导最近发现的p53同源物p73。至少当P73过量产生时，P73可以激活p53应答基因的转录，并通过诱导细胞凋亡来抑制细胞生长。由于内源性p73水平非常低，与p53不同，导致p73激活的上游信号通路尚未被阐明，因此E2F-1诱导p73水平显著增加是有趣的。E2F/pRb通路的失调在大多数肿瘤细胞中是常见的。因此，了解p73在这一途径中的功能可能为p73在肿瘤发生中的作用提供线索。特异性目的1的目的是确定E2F-1诱导p73的机制。将采用标准蛋白稳定性和定量RNA测定来确定E2F-1对p73的影响是否由于转录、mRNA稳定性和/或翻译后修饰的变化。E2F-1在p53失合细胞中诱导p73是否转化为p53靶基因的激活和凋亡将构成Specific Aims 2和3的基础。E2F-1诱导p53和p73靶基因的p73依赖性转激活的能力将在生化和基于转录的功能分析中进行分析。在Specific Aim 3中，流式细胞术和免疫荧光技术将用于研究p73是否参与E2F-1依赖性细胞凋亡。P73显性阴性蛋白将用于确定这些检测的特异性。最后，在Specific Aim 4中，我们将研究p73在化疗药物诱导的E2F依赖性细胞凋亡中的作用。这些研究可能为调控p73的信号提供了第一个线索，也可能有助于解释E2F对p53的独立杀伤。最终，本应用中提出的研究结果可能有助于开发新的基因疗法，其中E2F-1蛋白或p73蛋白可用于诱导肿瘤细胞凋亡。

项目成果

Vitamin D Receptor Activation Attenuates Hippo Pathway Effectors and Cell Survival in Metastatic Neuroblastoma.

• DOI: 10.1158/1541-7786.mcr-21-0425
• 发表时间: 2022-06-03
• 期刊: Molecular cancer research : MCR