PTEN TUMOR SUPPRESSOR AND SIGNAL TRANSDUCTION

描述：(改编自研究人员摘要)PTEN/MMAC1是一种肿瘤 在多种癌症中发生突变的抑制子。PTEN编码一种磷酸酶 认识到重要的第二信使， 磷脂酰肌醇-3，4，5-三磷酸(PIP3，4，5)，并去除 肌醇环上的3‘-磷酸。因此，PTEN可以对抗 磷脂酰肌醇(PI)-3激酶，它使肌醇环在 同样的立场。尽管PI-3激酶和PTEN潜在地影响许多 信号通路，可能是最被了解的通路物种，PTEN起作用 位于胰岛素受体和PI-3激酶下游，AKT上游。穿过 底物的磷酸化，AKT调节许多参与的靶点 转录、翻译、细胞周期和细胞凋亡。要测试 假设PTEN调节基因表达，他们开发了一种腺病毒 PTEN-/-肿瘤感染后2小时内表达PTEN的系统 细胞。PTEN的表达与AKT的快速抑制有关。已标记 来自不同感染时间的RNA与寡核苷酸芯片杂交 代表了超过40,000个基因。通过比较两种语言的表达水平 未感染的、腺-β-半乳糖苷酶和腺-PTEN感染的细胞，他们 确定了32名候选人，他们在3小时内至少被诱导了三次 感染。他们为24名候选人准备了探测器，其中8名 在Northern blotts上显示了PTEN特异性的诱导。最易被诱导的基因 是胰岛素受体底物(IRS)-2。IRS-2的诱导在 蛋白质水平。IRS-2是IRS基因家族中的一员，其功能是 胰岛素、胰岛素样生长因子和细胞因子受体传递的接头蛋白 信号。IRS-2的主要作用是将激活的受体连接到PI-3 激活剂。在PTEN感染的细胞中，诱导的IRS-2与 PI-3激酶。因此，他们的初步数据表明，PTEN的表达 诱导了一个反馈环来激活PI-3激酶。这项拨款申请将 探讨PTEN调控IRS-2的机制并定义 IRS-2在肿瘤发生中的作用他们将使用重组腺病毒和 用来识别细胞能够通过何种方式 诱导IRS2对PTEN的反应。这将包括使用腺病毒， 表达PTEN突变、显性阴性AKT和显性活性FKHR、GSK-3、 还有英国皇家空军。PI-3激酶、MEK和mTOR的抑制剂也将接受测试。他们 不仅会分析这些药物诱导IRS-2的能力，他们还会 同时比较它们与PTEN对AKT、MAPK、GSK-3和S6的影响 激活酶活性。PI-3激酶和AKT的p85和p110亚基是有效的 致癌基因。为了确定IRS-2在癌症中是否发生了基因改变，他们 将筛查细胞系和原发肿瘤中的突变和影响 含有野生型PTEN。因为IRS-2作为一种适配蛋白在 他们将研究PTEN对胰岛素信号转导的影响。 小鼠体内的胰岛素信号。最后，PTEN杂合子小鼠在 多个器官系统。他们将用IRS-2+/-小鼠培育PTEN+/-小鼠，以查看 IRS-2缺失是否能够减弱肿瘤表型。

DESCRIPTION: (Adapted from the investigator's abstract) PTEN/MMAC1 is a tumor suppressor that is mutated in many forms of cancer. PTEN encodes a phosphatase that recognizes the important second messenger, phosphatidylinositol-3,4,5-triphosphate (PIP3,4,5), and removes the 3'-phosphate from the inositiol ring. PTEN therefore antagonizes phosphatidylinositol (PI)-3 kinase, which phosphorylates the inositol ring at the same position. Although PI-3 kinase and PTEN potentially influence many signaling pathways, perhaps the best understood pathway species, PTEN acts downstream of the insulin receptor and PI-3 kinase and upstream of AKT. Through the phosphorylation of substrates, AKT regulates many targets involved in transcription, translation, the cell cycle, and apoptosis. To test the hypothesis that PTEN regulates gene expression, they developed an adenoviral system that expressed PTEN within two hours of infection in PTEN-/- tumor cells. PTEN expression was associated with a rapid inhibition of AKT. Labeled RNAs from different times of infection were hybridized to oligonucleotide chips representing over 40,000 genes. By comparing the level of expression between uninfected, Adeno-beta-galactosidase, and Adeno-PTEN infected cells, they identified 32 candidates that were induced at least threefold within 3 hours of infection. They prepared probes for 24 of the candidates, of which 8 demonstrated PTEN-specific induction on Northern blots. The most induced gene was the insulin receptor substrate (IRS)-2. Induction of IRS-2 was confirmed at the protein level. IRS-2 is a member of a family of IRS genes that function as adaptor proteins for the transmission of insulin, IGF, and cytokine receptor signals. The major role of IRS-2 is to link an activated receptor to PI-3 kinase. In PTEN infected cells, the induced IRS-2 coimmunoprecipitated with PI-3 kinase. Thus, their preliminary data suggests that the expression of PTEN induced a feedback-loop to activated PI-3 kinase. This grant application will attempt to explore the mechanism through which PTEN regulates IRS-2 and define the role of IRS-2 in tumorigenesis. They will use recombinant adenoviruses and pharmacological agents to identify the mode through which cells are able to induce IRS2 in response to PTEN. This will include the use of adenoviruses that express PTEN mutations, dominant-negative AKT and dominant active FKHR, GSK-3, and RAF. Inhibitors of PI-3 kinase, MEK, and mTOR will also be tested. They will not only analyze these agents for their ability to induce IRS-2, they will also compare them to PTEN regarding their effect on AKT, MAPK, GSK-3, and S6 kinase activity. The p85 and p110 subunits of PI-3 kinase and AKT are potent oncogenes. To determine whether IRS-2 is altered genetically in cancer they will screen for mutations and implications in cells lines and primary tumors containing wild type PTEN. Because IRS-2 functions as an adapter protein in the transduction of insulin signals, they will investigate the effect of PTEN on insulin signaling in mice. Finally, PTEN heterozygous mice have tumors in multiple organ systems. They will breed PTEN+/- mice with IRS-2+/- mice to see if IRS-2 loss is able to attenuate the tumor phenotype.