Dynamics, Regulation and Function of p53 in Single Cells

单细胞中 p53 的动态、调控和功能

• 批准号: 10321563
• 负责人: Galit Lahav
• 金额: $ 66.87万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-01-01 至 2025-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10321563
• 关键词: 3-Dimensional; Address; Affect; Architecture; Cell Death; Cells; Cellular Stress; Clinical; Conflict (Psychology); DNA Damage; Data; Development; Dose; Environment; Gene Combinations; Gene Expression; Genes; Goals; Human; Image; Immune; Immune system; Immunotherapy; Individual; Knowledge; Link; Malignant Neoplasms; Mediating; Normal Cell; Outcome; Pathway interactions; Pattern; Play; Population; Post-Translational Protein Processing; Process; Proteins; RNA; Radiation; Regulation; Role; Schedule; Signal Transduction; Stimulus; System; TP53 gene; Therapeutic; Time; Translating; Tumor Suppressor Proteins; Work; cancer cell; cancer therapy; cell killing; clinically relevant; combinatorial; imaging system; in vivo; inhibitor; insight; mutant; neoplastic cell; new technology; novel; overexpression; programs; response; single-cell RNA sequencing; targeted treatment; transcription factor; tumor

The dynamics of signaling systems are critical for controlling gene expression programs and cellular outcomes. The tumor suppressor protein p53 is a transcription factor orchestrating the response to cellular stresses, and we previously found that its dynamics (changes in its protein levels over time) following DNA damage depend on the stimulus and play a role in determining whether a cell will survive or die. However, many questions remain about how different cellular contexts influence p53 dynamics and ultimate cellular outcomes, how p53 chooses between conflicting cellular outcomes, and how p53 dynamics can best be leveraged for therapeutic purposes. The goals of our work are to obtain a comprehensive quantitative understanding of how p53 dynamics regulate cellular outcomes in single cells and to apply our findings to address clinical needs. The cellular environment can influence p53 dynamics, therefore we will first investigate how p53 dynamics are regulated by factors such as 3D cellular architecture in cultured tumor spheroids and in in vivo tumors. We will then investigate the dynamics and cellular outcomes of cancer-associated p53 mutants in cultured and in vivo settings. The effects of p53 dynamical patterns on gene expression will be determined in single cells by using novel technology that supports integrating live imaging data of p53 dynamics with single-cell RNA sequencing. We will also investigate how p53 dynamics influence gene expression at the RNA and protein levels, as well as the dynamics of p53 post-translational modifications in bulk populations. These studies will reveal the impact that p53 dynamical patterns have on the RNA and protein of its target genes, and how the combinations of these dynamical patterns guide cellular outcomes. We will also use our live-imaging systems to determine how clinically-relevant therapeutic approaches can be optimized to induce the desired p53 dynamics and cellular outcomes in cancer. We will determine how the doses and timings of radiation fractions affect p53 dynamics and function, and optimize the schedule of fractions for inducing tumor cell death via p53-mediated mechanisms. Many cancers overexpress the p53 inhibitors Mdm2 or Mdmx and are susceptible to their inhibition. Through quantifying and modulating p53 dynamics we will determine how to fine-tune their inhibition to sensitize Mdm2 or Mdmx overexpressing cells to DNA damage while sparing healthy cells. Tumor cells can be cleared by the immune system, and this process is influenced by the tumors' gene expression programs. Therefore, we will investigate how p53 dynamics influence interactions between tumor cells and immune cells, and work towards optimizing combinations of p53-targeting therapeutics with immunotherapies to maximize tumor cell killing by the immune system. In total, these studies will provide new mechanistic insights into the links between p53 dynamics and function in controlling cell fates, and will inform novel combinatorial therapeutic approaches to cancer treatments.

信号系统的动力学对于控制基因表达程序和细胞结果至关重要。 肿瘤抑制蛋白p53是一种转录因子，策划了对细胞应激的反应，并且 我们先前发现DNA损伤后其动力学（随着时间的推移随时间变化）取决于 在刺激上并在确定细胞是否生存还是死亡中发挥作用。但是，许多问题仍然存在 关于不同的细胞环境如何影响p53动力学和最终的细胞结果，p53如何选择 在相互冲突的细胞结局以及如何最好地利用p53动力学来治疗目的之间。 我们工作的目标是获得对p53动态如何调节的全面定量理解 单个细胞中的细胞结局并应用我们的发现以满足临床需求。细胞环境 可以影响p53动力学，因此我们将首先研究p53动态如何受到此类因素的调节 作为培养的肿瘤球体和体内肿瘤中的3D细胞结构。然后，我们将调查 培养和体内环境中与癌症相关的p53突变体的动力学和细胞结局。效果 通过使用新技术来确定基因表达上的p53动力学模式 支持通过单细胞RNA测序集成p53动力学的实时成像数据。我们还将调查 p53动力学如何影响RNA和蛋白质水平的基因表达以及p53的动力学 大量种群的翻译后修改。这些研究将揭示p53动力学的影响 模式具有其靶基因的RNA和蛋白质，以及这些动力学模式的组合如何 指导细胞结局。我们还将使用我们的现场模仿系统来确定临床上的相关性 可以优化治疗方法，以诱导癌症所需的p53动力学和细胞结局。 我们将确定辐射分数的剂量和时机如何影响p53动力学和功能，以及 优化通过p53介导的机制诱导肿瘤细胞死亡的分数时间表。许多癌症 过表达p53抑制剂MDM2或MDMX，并且容易受到抑制作用。通过量化和 调节p53动力学我们将确定如何微调其抑制作用以使MDM2或MDMX敏感 过表达细胞会导致DNA损伤，同时保留健康的细胞。可以通过免疫清除肿瘤细胞 系统，此过程受肿瘤的基因表达程序的影响。因此，我们将调查 p53动态如何影响肿瘤细胞与免疫细胞之间的相互作用，并致力于优化 p53靶向疗法与免疫疗法的组合，可通过免疫最大化肿瘤细胞杀死 系统。总的来说，这些研究将提供有关p53动力学与联系的新机械见解 在控制细胞命运方面的功能，并将为癌症治疗的新型组合治疗方法提供信息。