Spatiotemporal Regulation of Chk1 in Cell Biology, Cancer Etiology and Therapy

Chk1 在细胞生物学、癌症病因学和治疗中的时空调控

• 批准号: 8828117
• 负责人: YOU-WEI ZHANG
• 金额: $ 32.58万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-05-08 至 2016-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8828117
• 关键词: Attention; Binding; Binding Proteins; Biochemical; Biological Markers; Breast Cancer Patient; Breast Cancer cell line; Camptothecin; Cancer Etiology; Cancer cell line; Cell Culture Techniques; Cell Cycle Checkpoint; Cell Line; Cell Nucleus; Cell Survival; Cells; Cellular biology; Checkpoint kinase 1; Chemotherapy-Oncologic Procedure; Chromatin; Chromosomes; Complex; Cytoplasm; DNA; DNA Damage; DNA Repair; DNA-Binding Proteins; Data; Defect; Development; Diagnosis; Disease Progression; Embryonic Development; Event; Failure; Goals; Human; Human Genome; Immunohistochemistry; In Vitro; Knowledge; Laboratories; Laboratory Study; Lead; Leucine; Life; Light; MCM2 gene; MDA MB 231; Maintenance; Malignant Neoplasms; Measures; Modeling; Molecular; Mutagenesis; Nuclear; Nuclear Export; Pharmaceutical Preparations; Phase; Phosphorylation; Process; Progression-Free Survivals; Proteins; RNA Interference; Recruitment Activity; Regulation; Research; Resistance; Role; Signal Pathway; Signal Transduction; System; Testing; Tissues; Xenograft procedure; assault; cancer cell; cancer therapy; cell killing; chemotherapy; effective therapy; helicase; insight; mouse model; mutant; novel; novel strategies; predictive marker; repaired; response; spatiotemporal; surveillance network; transmission process; triple-negative invasive breast carcinoma; tumor; tumor growth; tumor initiation; ubiquitin ligase

DESCRIPTION (provided by applicant): DNA, the blueprint of life located inside our cells, is under continuous assault by both environmental and intracellular DNA damaging agents. If not corrected, these errors may lead to the development of cancer. Fortunately, cells have built up elegant DNA surveillance networks, termed cell cycle checkpoints, to detect and repair damaged DNA. Central to cell cycle checkpoints is a protein kinase, Chk1, which is essential for cell viability and mammalian embryonic development. Intriguingly, increasing evidence suggests that Chk1 may benefit tumor growth as well, as Chk1 expression is increased in malignant tumors and its level positively correlates with tumor grade. Most importantly, we recently showed that Chk1 is centrally involved in the resistance of cancer cells to chemotherapeutic drugs. These findings highlight the importance of Chk1 in cancer etiology and therapy. Therefore, a better understanding of Chk1 regulation not only provides insights the DNA damage response, but may also lead to the development of novel strategies in cancer therapy. Recent studies from this laboratory and others revealed an exciting spatiotemporal regulation model of Chk1, in which DNA damage induces phosphorylation and a rapid release of Chk1 from chromatin into the nucleus, and subsequently into the cytoplasm. Failure to undergo chromatin release of Chk1 impairs cell cycle checkpoints. On the other hand, cellular accumulation of active Chk1 proteins leads to the resistance of cancer cells to a clinically used chemotherapeutic drug, camptothecin (CPT). This is probably because Chk1-profient cells are better equipped to handle DNA damage induced by CPT. Thus, a general hypothesis is that the dynamic mobilization of Chk1 from one cellular compartment to another is crucial for the activation of cell cycle checkpoints in response to DNA damage, as well as for the chemotherapy response. In order to test this hypothesis, we first wish to understand the detailed molecular mechanisms underlying this chromatin->nucleus->cytoplasm mobilization of Chk1 in cells. We will first determine the factors that regulate chromatin association and disassociation of Chk1, a non-DNA binding protein (Aim 1). Second, we will define how Chk1 is exported from the nucleus to the cytoplasm (Aim 2). Further, we will interrogate the functional significance of the spatiotemporal regulation of Chk1. To do so, we will utilize a novel in vitro system established by this laboratory that highly recapitulates cell cycle checkpoints, as well as unique cell lines we generated with Chk1 mutant proteins specifically localized in a particular cellular compartment (Aim 2). Lastly, we will expand this study from a pure biochemical and molecular research into something that will offer novel insights into cancer etiology and chemotherapy through testing two hypotheses: one is that blocking the spatiotemporal mobilization of Chk1 sensitizes cancer cells to chemotherapeutic drugs, and the other is that Chk1 is a predictive marker for the therapy response of human tumors to chemotherapy (Aim 3).

描述(申请人提供)：DNA，位于我们细胞内的生命蓝图，正受到环境和细胞内DNA破坏因子的持续攻击。如果不加以纠正，这些错误可能会导致癌症的发展。幸运的是，细胞已经建立了优雅的DNA监视网络，称为细胞周期检查点，以检测和修复受损的DNA。细胞周期检查点的中心是一种蛋白激酶Chk1，它对细胞存活和哺乳动物胚胎发育至关重要。有趣的是，越来越多的证据表明，Chk1也可能有利于肿瘤的生长，因为Chk1在恶性肿瘤中的表达增加，其水平与肿瘤的分级呈正相关。最重要的是，我们最近发现Chk1与癌细胞对化疗药物的耐药性密切相关。这些发现突显了Chk1在癌症病因学和治疗中的重要性。因此，更好地了解Chk1的调控不仅有助于深入了解DNA损伤反应，而且可能有助于开发癌症治疗的新策略。本实验室和其他实验室最近的研究揭示了Chk1的一个令人兴奋的时空调控模型，在该模型中，DNA损伤导致Chk1从染色质中迅速磷酸化并释放到细胞核，然后进入细胞质。Chk1不能进行染色质释放会损害细胞周期检查点。另一方面，细胞内活性Chk1蛋白的积累导致癌细胞对临床使用的化疗药物喜树碱(CPT)产生耐药性。这可能是因为Chk1-profient细胞能够更好地处理CPT引起的DNA损伤。因此，一个普遍的假设是，Chk1从一个细胞室到另一个细胞室的动态动员对于激活细胞周期检查点以响应DNA损伤以及对化疗反应至关重要。为了验证这一假设，我们首先希望了解Chk1在细胞中的染色质-核-细胞质动员的详细分子机制。我们将首先确定调节染色质结合和解离的因素，Chk1是一种非DNA结合蛋白(目标1)。其次，我们将定义Chk1是如何从细胞核输出到细胞质的(目标2)。进一步，我们将询问Chk1时空调控的功能意义。为此，我们将利用本实验室建立的一种新的体外系统，该系统高度概括细胞周期检查点，以及我们用特定定位于特定细胞室的Chk1突变蛋白产生的独特细胞系(目标2)。最后，我们将把这项研究从纯粹的生化和分子研究扩展到通过检验两个假设来为癌症病因学和化疗提供新的见解：一个是阻断Chk1的时空动员使癌细胞对化疗药物敏感，另一个是Chk1是人类肿瘤对化疗的治疗反应的预测标记物(目标3)。