P16/P18 FAMILY CDK INHIBITORS

在大卫博士作为博士后研究员进行的先前研究中， 海滩在冷泉港实验室的实验室。发现 细胞周期蛋白-CDK复合物与许多小细胞 蛋白质，包括p2 l和p16。 P21（也称为CIP 1、WAF 1、SDII， CAP 20、PICI、CDKNI）通过至少三种途径调节细胞周期。 不同的途径：p2 l编码所有细胞周期蛋白CDK的有效抑制剂 p2 l蛋白与PCNA相互作用 独立于细胞周期蛋白-CDK酶抑制复制性DNA合成， p21也可以通过调节E2 F转录因子的活性， 它与细胞周期蛋白-CDK蛋白的相互作用。 p21的转录是 受肿瘤抑制因子p53的刺激，因此这种调节 提供了p53抑制肿瘤和细胞周期之间的直接联系 控制pl 6（也称为INK 4、MTSI、CDK 4 I、CDKN 2）编码细胞周期蛋白D- CDK 4特异性抑制剂，并且在多种肿瘤中缺失或突变。 人肿瘤源性细胞系和几种特定类型的原发性 肿瘤的 最近，P.I.的实验室发现了至少四种新的 CDK 4-和CDK 6-相互作用的小细胞蛋白质（pl 4、pl 5、pl 8和p20） 并已分离和功能性表征了编码以下两种的基因： 这四种蛋白，p18和p14 MTS 2。 pl 8和pl 4 MTS 2与 pl 6在序列、功能和进化中的作用，因此它们定义了一个新的 可能是CDK抑制剂的大家族。 四个具体目标 该建议涉及CbK抑制剂的p16/p18家族， 通过调节视网膜母细胞瘤基因的活性来控制细胞生长 产物pRb与人类的发育密切相关 癌 具体目标一。 新的CDK 4-和CDK 6-的鉴定和克隆 相关的细胞蛋白质。 本节中提出的实验是 目的在于鉴定和克隆编码新的CDK 4-和 CDK 6相关蛋白包括p15和p20。 具体目标二。 新的CDK 4-和CDK 6-相关的表征 细胞蛋白质。 本节中提出的实验旨在 编码CDK 4-和CDK 6-相关的基因的初步表征 通过特定目的I中提出的实验分离的蛋白质。 具体目标三。 pl 8和pl 6功能的机制。 中的实验 本节旨在（A）在体内证明 抑制途径，涉及p16/p18。(B)pRb-和 p53介导的P16转录抑制。 ~阐明 pl 6/pl 8抑制机制。 (D)p18在肌肉中的功能研究 细胞分化 具体目标四。寻找p18和p14 MTS 2突变。 在这 第一节，我们提出了寻找p18和MTS 2基因突变的实验 目的探讨p18抑瘤作用。

In previous studies conducted as a postdoctoral fellow in Dr. David Beach's laboratory at Cold Spring Harbor Laboratory, the P.I. discovered that cyclin-CDK complexes are associated with a number of small cellular proteins, including p2l and pl6. P2l (also known as CIPI, WAF1, SDII, CAP20, PICI, CDKNI) regulates the cell cycle through at least three different pathways: p2l encodes an potent inhibitor of all cyclin CDK enzymes that have been tested, p2l protein interacts with PCNA independently of cyclin-CDK enzymes to inhibit replicative DNA synthesis, and p2l can also regulate the activity of the E2F transcription factor via its interaction with cyclin-CDK proteins. Transcription of p21 is stimulated by the tumor suppressor p53, and therefore this regulation provides a direct link between tumor suppression by p53 and cell cycle control. pl6 (also known as INK4, MTSI, CDK4I, CDKN2) encodes a cyclin D- CDK4 specific inhibitor and is deleted or mutated in a wide variety of human tumor-derived cell lines and in several specific types of primary tumors. Very recently, the P.I.'s laboratory identified at least four new CDK4- and CDK6-interacting small cellular proteins (pl4, pl5, pl8 and p20) and have isolated and functional characterized the gene encoding two of these four proteins, pl8, and p14MTS2. pl8 and pl4MTS2 are related to pl6 in sequence, function and evolution, and thus they define a new and potentially large family of CDK inhibitors. The four specific aims of this proposal are concerned with the pl6/pl8 family of CbK inhibitors that controls cell growth by regulating the activity of the retinoblastoma gene product, pRb, and that is strongly implicated in the development of human cancer. Specific Aim I. Identification and Cloning of Novel CDK4- and CDK6- Associated Cellular Proteins. Experiments proposed in this section are aimed at identification and cloning of genes encoding novel CDK4- and CDK6-associated proteins including pl5 and p20. Specific Aim II. Characterization of Novel CDK4- and CDK6-Associated Cellular Proteins. Experiments proposed in this section aim at the initial characterization of genes encoding CDK4-and CDK6-associated proteins isolated by the experiments proposed in Specific Aim I. Specific Aim III. Mechanism of pl8 and pl6 Function. The experiments in this section are aimed at (A) Demonstration in vivo of a growth suppression pathway involving pl6/pl8. (B) Characterization of pRb- and p53-mediated repression of pl6 transcription. ~ Elucidation of the pl6/pl8 inhibitory mechanism. (D) Investigation of p18 function in muscle cell differentiation. Specific Aim IV. Searching for p18 and p 14MTS2 mutations. In this section, we propose experiments to search for p18 and MTS2 gene mutations t investigate p18 tumor suppression function.