Role of CDK4 in Cell Cycle Progression and Cancer

描述（由申请人提供）：细胞周期蛋白依赖性激酶4（Cdk 4）是一种 哺乳动物细胞G1/S细胞周期进程重要调节因子 文化该基因第24位密码子的种系突变（R至C）导致 个体发展为黑素瘤的倾向。到 为了了解这种酶在体内的作用，我们靶向了小鼠cdk 4 通过胚胎干细胞和产生的菌株中的同源重组获得基因座 缺乏Cdk 4表达（Cdk 4 neo/neo）或表达激活的 这种酶的一种形式（Cdk 4 R24 C/R24 C），不能与细胞周期蛋白相互作用。 激酶抑制剂p161 NK 4A。纯合子Cdk 4（Cdk 4 neo/neo）无效突变小鼠是 有活力，但在生长、精子发生和卵子发生方面表现出缺陷。在 此外，发现这些小鼠是糖尿病小鼠，其表现出有缺陷的胰腺炎。 β细胞发育、多尿和多饮。与此相反，Cdk 4 R24 C/R24 C 小鼠表现出生长优势和胰腺β细胞增生， 类似胰岛素瘤。此外，这些小鼠自发地发展出肿瘤， 不同的细胞来源，清楚地表明了解除管制的后果， cdk 4介导的细胞通路在各种类型的癌症的进展。 为了进一步了解Cdk 4在细胞周期调控中的作用， 肿瘤，我们建议：（1）进行详细的表征MEFs和 来自cdk 4（R24 C/R24 C）小鼠的淋巴细胞，以确定R24 C的作用 细胞周期动力学的突变，胚胎成纤维细胞经历 衰老，成纤维细胞和淋巴细胞对凋亡刺激的敏感性; 以及MEFs对癌基因引起的肿瘤转化的易感性。(2)到 将目标1中描述的实验扩展到整个动物模型系统， 我们的实验旨在确定增强 Cdk 4 R24 C/R24 C小鼠对癌发展的易感性 在用化学致癌物如TPA和/或DMBA处理之后。(3)到 研究Cdk 4-pRb和p53通路之间的协作，使用 Cdk 4 neo/neo和Cdk 4 R24 C/R24 C小鼠和p53缺陷小鼠。(4)测试 ras激活和pRb失活的协同作用， 携带两种突变的小鼠的肿瘤发展特征， 将Cdk 4 R24 C/R24 C小鼠与携带活化生殖系的小鼠杂交 ras基因突变。

DESCRIPTION (provided by applicant): Cyclin-dependent kinase 4 (Cdk4) is an important regulator of G1/S cell cycle progression of mammalian cells in culture. Germline mutations in the 24th codon of this gene (R to C) results in a predisposition of the individuals to the development of melanoma. To understand the role of this enzyme in vivo, we have targeted the mouse cdk4 locus by homologous recombination in embryonic stem cells and generated strains of mice that either lack Cdk4 expression (Cdk4 neo/neo) or express an activated form of this enzyme (Cdk4 R24C/R24C), which cannot interact with the Cyclin Kinase Inhibitor p161NK4A. Homozygous Cdk4 (Cdk4 neo/neo) null mutant mice are viable but express defects in growth, spermatogenesis and oogenesis. In addition, these mice were found to be diabetic exhibiting defective pancreatic beta-cell development, polyuria and polydypsia. In contrast, Cdk4 R24C/R24C mice exhibit a growth advantage, and pancreatic beta-cell hyperplasia which resembled insulinomas. Moreover, these mice spontaneously develop tumors of varying cellular origin, clearly demonstrating the consequence of de-regulated Cdk4 mediated cellular pathways in the progression of various types of cancer. To gain further insight into the role of Cdk4 in cell cycle regulation and neoplasia, we propose: (1) To carry out a detailed characterization of MEFs and lymphocytes derived from cdk4 (R24C/R24C) mice to determine the effect of R24C mutation on cell cycle kinetics, ability of embryonic fibroblasts to undergo senescence, susceptibility of fibroblasts and lymphocytes to apoptotic stimuli; and susceptibility of MEFs to neoplastic transformation by oncogenes. (2) To extend experiments described in Aim 1 to whole animal model systems by carrying out experiments aimed at determining the molecular basis for the enhanced susceptibility of the Cdk4 R24C/R24C mice to the development of carcinomas following treatment with chemical carcinogens such as TPA and/or DMBA. (3) To investigate the collaboration between the Cdk4-pRb and p53 pathways, using Cdk4neo/neo and Cdk4R24C/R24C mice and mice deficient in p53. (4) To test the co-operating effects of ras activation and pRb inactivation by examining the tumor development characteristics of mice that harbor both mutations by crossing Cdk4R24C/R24C mice with mice that carry an activating germline mutation in the ras gene.