FANCJ dependent pathways in replication stress

复制应激中的FANCJ依赖性途径

基本信息

批准号：
10219989
负责人：
Sharon B Cantor
金额：
$ 35.87万
依托单位：
UNIV OF MASSACHUSETTS MED SCH WORCESTER
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-09-05 至 2023-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10219989
关键词：
Acetylation Address BRCA1 gene BRCA2 gene Binding Biochemical Genetics Bioinformatics Biological Assay Breast Breast Cancer gene Bypass Cancer Patient Cell Proliferation Cell Survival Cells Cisplatin Clinical Complex DNA DNA Damage DNA Mismatch Repair Protein MLH1 DNA Repair DNA Repair Gene DNA Structure DNA biosynthesis DNA lesion DNA replication fork Data Defect Disease Electron Microscopy Engineering Ensure Equilibrium Fanconi Anemia-BRCA Pathway Fanconi anemia protein Fanconi&apos s Anemia Fiber Functional disorder Genes Genome Genomic Instability Goals Health Hereditary Breast Carcinoma Hereditary Breast and Ovarian Cancer Syndrome Knowledge Link MLH1 gene Malignant Neoplasms Malignant neoplasm of ovary Mismatch Repair Modification Molecular Mutate Mutation Null Lymphocytes Oncogenes Outcome Pathway interactions Patients Phenotype Phosphorylation Play Process Proteins Proteomics Recovery Regulation Research Resistance Role SMARCA3 gene Site Stress Supporting Cell Testing Therapeutic Therapeutic Intervention Transitional Cell Translations Tumor Suppression biological adaptation to stress chemotherapy gain of function gene function genetic approach genome integrity helicase improved innovation insight mutant repaired replication stress single molecule therapeutic development transcription factor tumor

项目摘要

Project Summary: Great progress has been made in uncovering the proteins and pathways that function in the replication stress response. In particular, the hereditary breast cancer genes, as well as genes mutated in Fanconi anemia (FA) function in the replication stress response. It is now understood that loss of their function in the replication stress response contributes to the sensitivity of associated tumors to chemotherapies, such as cisplatin. However, the distinct functions for the BRCA-FA proteins are largely unknown. Here, we propose to analyze DNA replication fork dynamics, replisome components, and identify patient mutations that have specific defects in the replication stress response. To define how a cell transitions from defective to dysregulated replication, we have engineered cells expressing different mutant versions of the BRCA1-associated FANCJ also mutated in breast/ovarian cancer and FA. Similar to BRCA1, we have uncovered that FANCJ functions to protect replication forks from collapse. We also found that this FANCJ fork protection function requires its direct interaction with the mismatch repair (MMR) protein, MLH1. This finding provides insight as to why cells lacking the FANCJ-MLH1 interaction fail to recover from replication stress. We have also identified putative gain-of-function FANCJ mutants, such as the BRCA1-interaction defective mutant, that circumvent replication stress, keep forks intact, and confer hyper- resistance to replication stress inducing agents. In Aim 1, we will seek to define how FANCJ interactions direct DNA replication fork dynamics. Given that FANCJ localizes to replication forks, displaces proteins, and unwinds DNA, we hypothesize that disrupted vs dysregulated replication will reflect not only changes in DNA structures, but also the proteins found at DNA replication forks. In Aim 2, we will seek to determine how FANCJ contributes to the composition of the replisome in both unchallenged and at stressed replication forks. Replication stress induces changes to FANCJ protein interactions and post-translation modifications. Some of these changes occur at sites we found to be mutated in cancer patients. In Aim 3, we will seek to generate FANCJ mutants resistant to replication stress induced changes to uncover mechanisms regulating FANCJ function that are lost in cancer. Collectively, by defining how cells succumb to- or survive- toxic DNA damage that normally interferes with replication, we will gain insight towards mechanisms transitioning cells from defective to dysregulated replication in cancer. !

项目概要：在揭示复制应激中发挥作用的蛋白质和途径方面取得了巨大进展回复。特别是遗传性乳腺癌基因以及范可尼贫血 (FA) 突变基因在复制应激反应中发挥作用。现在人们知道，它们在复制过程中功能的丧失应激反应有助于相关肿瘤对化疗（例如顺铂）的敏感性。然而，BRCA-FA 蛋白的独特功能很大程度上未知。在此，我们建议分析一下 DNA 复制叉动力学、复制体成分，并识别具有特定缺陷的患者突变在复制应激反应中。为了定义细胞如何从复制缺陷转变为复制失调，我们设计了细胞表达 BRCA1 相关 FANCJ 的不同突变版本也在乳腺癌/卵巢癌中发生突变和FA。与 BRCA1 类似，我们发现 FANCJ 具有保护复制叉免于崩溃的功能。我们还发现FANCJ叉子保护功能需要与失配修复直接交互 (MMR) 蛋白，MLH1。这一发现提供了关于为什么缺乏 FANCJ-MLH1 相互作用的细胞无法从复制压力中恢复。我们还鉴定了假定的功能获得性 FANCJ 突变体，例如 BRCA1 相互作用缺陷突变体，可规避复制压力，保持分叉完整，并赋予超对复制应激诱导剂的抗性。在目标 1 中，我们将寻求定义 FANCJ 交互如何指导 DNA 复制叉动力学。鉴于 FANCJ 定位于复制叉，置换蛋白质，并且解开 DNA，我们假设复制中断与失调不仅反映 DNA 的变化结构，还有 DNA 复制叉上发现的蛋白质。在目标 2 中，我们将寻求确定如何 FANCJ 有助于未受挑战和受应激复制叉中复制体的组成。复制压力会引起 FANCJ 蛋白相互作用和翻译后修饰的变化。一些这些变化发生在我们发现癌症患者发生突变的位点。在目标 3 中，我们将寻求产生抵抗复制应激的 FANCJ 突变体诱导的变化揭示了调节 FANCJ 的机制癌症中丧失的功能。总的来说，通过定义细胞如何屈服于有毒 DNA 损伤或存活下来通常会干扰复制，我们将深入了解细胞从癌症中复制失调的缺陷。！