Characterization of JNK in Cell Cycle Control

JNK 在细胞周期控制中的表征

• 批准号: 7676182
• 负责人: Ze'ev A Ronai
• 金额: $ 39.63万
• 依托单位: SANFORD BURNHAM PREBYS MEDICAL DISCOVERY INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-08-19 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7676182
• 关键词: Address; Affect; Animal Model; Biochemical; Boxing; Cell Cycle; Cell Cycle Arrest; Cell Cycle Checkpoint; Cell Cycle Progression; Cell Cycle Regulation; Cell Death; Cells; Cellular Stress Response; Cellular biology; Chromosomal Instability; Collaborations; Cultured Cells; Cyclin B; DNA Damage; DNA damage checkpoint; Data; Equilibrium; Exhibits; Family; Family member; Foundations; G1 Phase; G2/M Arrest; Generations; Genetic; Genetic Models; Genotoxic Stress; Human; Human Development; In Vitro; JNK-activating protein kinase; Knock-in Mouse; Link; Lobe; MAPK8 gene; MAPK9 gene; Malignant Neoplasms; Mammalian Cell; Mediating; Meiosis; Microtubules; Mitosis; Mitotic spindle; Modeling; Molecular; Mus; Mutant Strains Mice; Oocytes; Pathway interactions; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphotransferases; Physiological; Play; Protein Isoforms; Protein Kinase; Proteins; Recruitment Activity; Regulation; Role; Scaffolding Protein; Stress; System; Testing; Ubiquitination; Xenopus laevis; anaphase-promoting complex; base; biological adaptation to stress; cytokine; genetic analysis; genetic regulatory protein; in vivo; melanoma; member; mouse model; multicatalytic endopeptidase complex; novel; public health relevance; reconstitution; response; segregation; stress activated protein kinase; tumor; ubiquitin ligase; ultraviolet irradiation

DESCRIPTION (provided by applicant): JNK (1-3) are members of the stress-activated protein kinase family that regulates the cellular stress response via their control of cell death and survival. Our recent studies disclose an unexpected mechanism for the regulation and consequently the function of JNK; our preliminary results demonstrate that through its KEN-box, JNK is targeted for degradation in a cell cycle-dependent manner. Such targeting is mediated by Cdh1, which recruits the Anaphase Promoting Complex or Cyclosome (APC/C) ubiquitin ligase to ubiquitinate JNK, resulting in its proteasome-dependent degradation. APC/CCdh1-mediated JNK degradation primarily occurs during the exit from mitosis, and G1 phases of the cell cycle. Our preliminary data also identify Cdc25C as JNK substrate at the G2/M-G1 phases of the cell cycle. JNK degradation is important for the regulation of Cdc25C activity and Wee1 stability. Thus, a non-degradable (yet which can be activated) form of JNK (JNKKEN) induces reduced cyclin-B/Cdk1 activity, affects spindle and chromosomal dynamics and delays exit from mitosis. These findings provide the foundation for our hypothesis that through the regulation of Cdc25C, JNK plays an important role in control of cell cycle progression during which it is tightly regulated by the APC/CCdh1. Our finding that JNK is a cell cycle regulator offers an undisclosed link between JNK and unrestrained cell cycle progression as well as chromosome instability, commonly observed in human cancers, where the balance between JNK functions as a stress kinase and cell cycle control protein might be altered. To test our hypothesis we propose to carry out the following studies: Aim 1 - Assess the molecular determinants responsible for JNK activity during the cell cycle and determine which of JNK family members is most important for cell cycle control.; Aim 2- characterize the effect of JNK on Cdc25C as part of its control of cell cycle transition. Aim 3 -Characterize the cytological changes in mitosis that are controlled by JNK and Cdc25C regulation by JNK; Aim 4 - determine the physiological significance of JNK's role in cell cycle control in a JNK-KEN knock-in mouse model. Aim 5 - using genetic, biochemical and cell biology approaches, determine the role of JNK in cell cycle checkpoint pathways following stress and DNA damage and in human melanoma as a model for constitutively deregulated JNK activity. This proposal brings together experts that closely collaborate to address an important and novel function of the key kinase, JNK. This concerted effort will provide the foundation for understanding the role of JNK in cell cycle progression and the implication of its deregulated control to genotoxic stress response as well as to development of human cancer. PUBLIC HEALTH RELEVANCE: This application will test the hypothesis that JNK is important player in cell cycle control prior and more so, in response to DNA damage. Support for this hypothesis comes from studies in which we discovered that JNK is targeted for degradation at G2M-G1 phases of the cell cycle. JNK mediates its function through phosphorylation of Cdc25C which affects its phosphatase activity. Interfering with JNK degradation or altering its activity delays exit from mitosis and impairs G2 arrest after DNA damage. Using biochemical, cellular and genetic models our proposed studies will delineate the requirements and regulation of this newly identified regulatory lobe along the cell cycle and DNA damage response.

描述（由申请人提供）：JNK（1-3）是应激激活蛋白激酶家族的成员，通过控制细胞死亡和存活来调节细胞应激反应。我们最近的研究揭示了一种意想不到的调控机制，从而揭示了JNK的功能；我们的初步结果表明，通过其KEN-box， JNK以细胞周期依赖的方式被降解。这种靶向是由Cdh1介导的，Cdh1招募后期促进复合物或环体（APC/C）泛素连接酶使JNK泛素化，导致其蛋白酶体依赖性降解。APC/ ccdh1介导的JNK降解主要发生在有丝分裂结束和细胞周期的G1期。我们的初步数据还确定Cdc25C在细胞周期的G2/M-G1期作为JNK底物。JNK降解对Cdc25C活性和Wee1稳定性的调控很重要。因此，JNK （JNKKEN）的不可降解（但可以激活）形式诱导细胞周期蛋白b /Cdk1活性降低，影响纺锤体和染色体动力学并延迟有丝分裂的退出。这些发现为我们的假设提供了基础，即通过调控Cdc25C， JNK在细胞周期进程中发挥重要作用，在此过程中它受到APC/CCdh1的严格调控。我们发现JNK是一种细胞周期调节剂，提供了JNK与不受约束的细胞周期进程以及染色体不稳定性之间未公开的联系，在人类癌症中通常观察到，其中JNK作为应激激酶和细胞周期控制蛋白之间的平衡可能被改变。为了验证我们的假设，我们建议进行以下研究：目的1 -评估在细胞周期中负责JNK活性的分子决定因素，并确定JNK家族成员中哪些对细胞周期控制最重要。目的2-表征JNK对Cdc25C的作用，作为其控制细胞周期转变的一部分。目的3 -表征JNK和Cdc25C调控的有丝分裂细胞学变化；目的4 -在JNK- ken敲入小鼠模型中确定JNK在细胞周期控制中的生理意义。目的5 -利用遗传、生化和细胞生物学方法，确定JNK在应激和DNA损伤后的细胞周期检查点途径中的作用，并在人类黑色素瘤中作为JNK活性组成性失调的模型。该提案汇集了密切合作的专家，以解决关键激酶JNK的重要和新功能。这一共同努力将为理解JNK在细胞周期进程中的作用以及它对基因毒性应激反应和人类癌症发展的不受管制的影响提供基础。公共卫生相关性：该应用将验证JNK在细胞周期控制中起重要作用的假设，在DNA损伤反应中更是如此。我们在研究中发现，JNK在细胞周期的G2M-G1期是降解的目标，这些研究支持了这一假设。JNK通过磷酸化Cdc25C介导其功能，从而影响其磷酸酶活性。干扰JNK降解或改变其活性会延迟有丝分裂的退出，并损害DNA损伤后的G2阻滞。利用生物化学、细胞和遗传模型，我们提出的研究将沿着细胞周期和DNA损伤反应描绘这个新发现的调节叶的要求和调节。