Chemical Genetic Analysis of the Human Cell Cycle

人类细胞周期的化学遗传分析

基本信息

批准号：
8727082
负责人：
ROBERT P FISHER
金额：
$ 32.07万
依托单位：
ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-09-01 至 2017-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8727082
关键词：
Ablation Active Sites Alleles Antineoplastic Agents Apoptosis Benefits and Risks Binding Binding Sites Biological Process Cell Cycle Cell Cycle Regulation Cell Fraction Cell Proliferation Cell Proliferation Regulation Cell division Cells Cessation of life Checkpoint kinase 1 Colon Carcinoma Complement Complex Coupled Cyclin-Dependent Kinase Inhibitor Cyclin-Dependent Kinases Cyclins DNA Damage DNA Double Strand Break DNA biosynthesis Development Discrimination Dissection Ensure Enzymes Event Family Feedback Financial compensation Gene Expression Genes Genetic Techniques Genomics Goals Growth and Development function Human Human Engineering Individual Ionizing radiation Logic Malignant Neoplasms Mammalian Cell Mitogens Mus Mutate Mutation Nijmegen Breakage Syndrome Normal tissue morphology Organism Pathway interactions Phosphotransferases Physiological Population Proteins Regulation Role S Phase Specificity Stress Testing Time Toxic effect Work adenine analog cancer cell cell killing cell type chemical genetics chemotherapy cyclin-dependent kinase-activating kinase falls flexibility gain of function genetic analysis inhibitor/antagonist insight knockout gene novel strategies premature prevent public health relevance repaired response scaffold small molecule tumor

项目摘要

DESCRIPTION (provided by applicant): Proper regulation of cell division is required for normal growth, development and genomic integrity; aspects of this control are lost or disrupted in human cancers. The cyclin-dependent kinases (CDKs) drive all major transitions in the cell division cycle. Mammalian cells have multiple downstream effector CDKs but, as we show, a single upstream CDK-activating kinase (CAK), Cdk7. Gene knockout and silencing studies suggested redundancy in the CDK network, by showing that Cdk2-long thought to be the principal driver of DNA synthesis (S) phase-was dispensable for viability. However, our work-using a chemical-genetic approach that selectively inactivates catalytic function of the targeted CDK while sparing non-catalytic, "scaffold" functions-uncovered strict requirements for Cdk2 activity in proliferation of both transformed and non- transformed human cells. An important function of Cdk2 is to prevent premature activation of Cdk1, and thereby ensure coordinated progression through S phase. That function depends on a kinetically more favorable pathway for activation of Cdk2, compared to Cdk1, due in part to different mechanisms of recognition by Cdk7. I hypothesize that specialized functions of Cdk2-in regulating gene expression, coordinating S phase and responding to DNA damage or replication stress-emerge from its unique mode of activation. I propose, moreover, that another distinct pathway by which Cdk7 activates Cdk4 and Cdk6-which function prior to cell-cycle commitment-is directly or indirectly coupled to mitogen-sensing pathways. The specific aims are: 1. To dissect the G1/S regulatory network comprising Cdk7, Cdk2, Cdk4 and Cdk6 2. To probe specific functions of Cdk7 and Cdk2 in the DNA damage response 3. To target the Cdk2 activation pathway with small molecule inhibitors Our preliminary studies reveal a unitary CAK-CDK network that achieves regulatory flexibility through kinetically distinct modes of kinase activation and inactivation. I ow propose to probe specific functions of CAK and the downstream CDKs in coordinating constitutive cell-cycle events and ensuring effective responses to genotoxic insults; and to test a new paradigm for inhibiting a specific CDK by selective targeting of its activation pathway. These studies promise to advance basic understanding of cell-cycle control, and to reveal novel strategies for anti-CDK therapy of human cancer.

描述（由申请人提供）：正常生长，发育和基因组完整性需要适当的细胞分裂调节；这种控制的各个方面在人类癌症中丢失或破坏。依赖细胞周期蛋白的激酶（CDKS）驱动细胞分裂周期中的所有主要过渡。哺乳动物细胞具有多个下游效应子CDK，但是如我们所示，是一个上游CDK激活激酶（CAK），CDK7。基因敲除和沉默研究表明，CDK网络中的冗余性，表明CDK2长被认为是DNA合成的主要驱动力（S）相 - WAS-WAS可用于生存能力。但是，我们使用的工作遗传方法有选择地使靶向CDK的催化功能有选择地失活，同时避免了非催化性，“支架”功能 - 对CDK2活性的严格要求在转化和非转化的人类细胞的增殖中。 CDK2的重要功能是防止CDK1的过早激活，从而确保通过S相协调进展。与CDK1相比，该功能取决于动力学上更有利的CDK2激活途径，部分原因是CDK7的识别机制不同。我假设CDK2-IN调节基因表达的专门功能，协调S相并从其独特的激活方式响应DNA损伤或复制应力发出。此外，我提出，CDK7激活CDK4和CDK6的另一种不同的途径 - 在细胞周期投入之前的功能直接或间接与有丝分裂原敏感途径耦合。 The specific aims are: 1. To dissect the G1/S regulatory network comprising Cdk7, Cdk2, Cdk4 and Cdk6 2. To probe specific functions of Cdk7 and Cdk2 in the DNA damage response 3. To target the Cdk2 activation pathway with small molecule inhibitors Our preliminary studies reveal a unitary CAK-CDK network that achieves regulatory flexibility through kinetically distinct modes of kinase激活和灭活。我建议在协调组成型细胞周期事件中探测CAK和下游CDK的特定功能，并确保对遗传毒性损伤的有效反应；并测试通过选择性靶向其激活途径来抑制特定CDK的新范式。这些研究有望提高对细胞周期控制的基本了解，并揭示针对人类癌症抗CDK治疗的新策略。