Role of nuclear profilin-1 in DNA replication fork stability and cancer chemotherapy response

核 profilin-1 在 DNA 复制叉稳定性和癌症化疗反应中的作用

• 批准号: 10587921
• 负责人: Jieya Shao
• 金额: $ 39.3万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-12-01 至 2028-02-29
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10587921
• 关键词: Actin-Binding Protein; Actins; Binding; Breast Cancer Cell; Camptothecin; Cell Death; Cell Nucleus; Cells; Characteristics; Chemoresistance; Chemotherapy-Oncologic Procedure; Chromatin; Chromosome abnormality; Clinical; Complex; Cytoplasm; DNA; DNA Damage; DNA Maintenance; DNA Repair; DNA biosynthesis; DNA lesion; DNA replication fork; Data; Enzymes; Equilibrium; Excision; Exportins; FANCD2 protein; Foundations; Funding; Genomic Instability; Grant; Histones; Hypersensitivity; Investigation; Knowledge; Malignant Neoplasms; Mammalian Cell; Methylation; Modeling; Molecular; Names; Natural Immunity; Nuclear; Nucleosomes; Oncogenes; Outcome; Phenotype; Play; Poly(ADP-ribose) Polymerase Inhibitor; Polymers; Predisposition; Process; Proline; Proteins; Publishing; Rationalization; Regulation; Relaxation; Reporting; Role; SMARCA5 gene; Single-Stranded DNA; Slide; Solid; Source; Stimulator of Interferon Genes; Stress; Structure; Testing; Therapeutic; Time; Transcription Elongation; Tumor Suppressor Proteins; Up-Regulation; War; Work; anti-cancer; anticancer activity; arm; brca gene; cancer cell; chemotherapy; cofactor; coping mechanism; genotoxicity; hydroxyurea; insight; micronucleus; novel; nuclease; polymerization; profilin 1; replication stress; stem; treatment response; tumor; tumor growth

PROJECT SUMMARY DNA replication stress is one of the mechanistic underpinnings of many genotoxic chemotherapies which cause replication fork stalling. Stalled forks frequently reverse to form four-way junction structures, as a natural coping mechanism, to allow repair of the DNA lesions ahead of the forks and avoid catastrophic fork collapse. However, fork reversal is a double-edged sword because the nascent DNA in the reversed arm is susceptible to excessive nuclease resection if not securely protected which can lead to severe genome instability. Many factors play tug- of-war at stalled forks to strengthen or weaken their stability, and their balance in the cell determines the fate of replication forks during stress and cellular outcome upon chemotherapy treatments. In this study, we investigate the molecular mechanisms of a novel replication fork regulator named profilin-1 (Pfn1). As a well-known actin- binding protein, Pfn1 plays an essential role in actin polymerization and dynamics. Paradoxically, it also has well- documented but poorly understood anticancer activities including the ability to sensitize cancer cells to chemotherapy treatments. In recently published work, we demonstrated for the first time that the anticancer effects of Pfn1 stem, at least partially, from its nuclear functions that are spatially and mechanistically distinct from its cytoplasmic function in actin regulation. We showed that nuclear Pfn1 directly interacts with ENL in the Super Elongation Complex (SEC) and inhibits the ability of SEC to drive transcriptional elongation of various cancer genes including MYC. We also presented clinical evidence that nuclear Pfn1 level is frequently decreased in cancer due to the upregulation of its nuclear exporter exportin-6 (XPO6), whose deletion increases nuclear Pfn1 level and decreases tumor growth. These findings establish the notion that Pfn1 has fundamentally important and cancer-relevant functions in the nucleus, and set the stage for further discovery of its involvement in additional nuclear processes. In this grant, we present novel evidence that nuclear Pfn1 promotes normal DNA replication but causes fork destabilization during stress and increases cellular sensitivity to replication stress- inducing chemotherapies including PARP inhibitors. We hypothesize that nuclear Pfn1 has context-dependent effects on DNA replication forks. Under normal conditions, it promotes fork progression by increasing chromatin relaxation through SNF2H. Under stressed conditions, it promotes fork reversal by stimulating SNF2H and increases fork resection by suppressing BOD1L, leading to fork destabilization. Aim 1: Determine the effect of nuclear Pfn1 on BOD1L-dependent replication fork protection. Aim 2: Define the role of Pfn1/SNF2H axis in replication fork remodeling and stability. Aim 3: Understand the chemotherapy-sensitizing ability of nuclear Pfn1. Work proposed in this grant has the potential to generate important mechanistic insights and proof-of-concept therapeutic data rationalizing further investigation of nuclear Pfn1 as a synthetic lethal target for genotoxic chemotherapies.

项目概要 DNA 复制应激是许多基因毒性化疗的机制基础之一，可导致 复制叉停滞。停滞的叉子经常反转形成四向连接结构，作为一种自然的应对方式 机制，以修复分叉之前的 DNA 损伤，避免灾难性的分叉崩溃。然而， 叉逆转是一把双刃剑，因为逆转臂中的新生 DNA 很容易受到过度的影响。 核酸酶切除如果没有得到安全保护，可能会导致严重的基因组不稳定。诸多因素发挥着拉扯作用—— 停滞的叉子上的战争会增强或削弱它们的稳定性，它们在细胞中的平衡决定了叉子的命运 在化疗治疗的压力和细胞结果期间复制分叉。在这项研究中，我们调查 名为 profilin-1 (Pfn1) 的新型复制叉调节因子的分子机制。作为知名的行动者 结合蛋白 Pfn1 在肌动蛋白聚合和动力学中起着重要作用。矛盾的是，它也有很好的—— 有记录但知之甚少的抗癌活​​性，包括使癌细胞敏感的能力 化疗治疗。在最近发表的工作中，我们首次证明了抗癌药物 Pfn1 的作用至少部分源于其在空间和机制上不同的核功能 来自其在肌动蛋白调节中的细胞质功能。我们发现核 Pfn1 直接与 ENL 相互作用 超级延伸复合物 (SEC) 并抑制 SEC 驱动各种转录延伸的能力 癌症基因包括 MYC。我们还提供了核 Pfn1 水平经常降低的临床证据 由于其核输出蛋白exportin-6 (XPO6) 的上调，该蛋白的缺失增加了核 Pfn1 水平并减少肿瘤生长。这些发现证实了 Pfn1 从根本上具有 细胞核中重要且与癌症相关的功能，并为进一步发现其参与奠定了基础 在额外的核过程中。在这笔资助中，我们提出了核 Pfn1 促进正常 DNA 的新证据 复制，但会在压力期间导致分叉不稳定，并增加细胞对复制压力的敏感性 诱导化疗，包括 PARP 抑制剂。我们假设核 Pfn1 具有上下文依赖性 对 DNA 复制叉的影响。在正常情况下，它通过增加染色质来促进分叉进展 通过 SNF2H 放松。在压力条件下，它通过刺激 SNF2H 和 通过抑制 BOD1L 增加叉切除，导致叉不稳定。目标 1：确定效果 核 Pfn1 对 BOD1L 依赖性复制叉的保护。目标 2：定义 Pfn1/SNF2H 轴在 复制叉重塑和稳定性。目标 3：了解核 Pfn1 的化疗增敏能力。 这笔赠款中提出的工作有可能产生重要的机制见解和概念验证 治疗数据合理化进一步研究核 Pfn1 作为基因毒性合成致死靶点 化疗。