Phosphoinositide Signaling in the Cytosol and Nucleus

项目摘要。本研究旨在了解空间磷酸肌醇信号（PI）机制 在细胞质和细胞核中。这些通路对癌症、神经发育障碍、 糖尿病和几种先天性疾病。在细胞质中，激动剂，如EGF（和许多其他），激活 控制大多数细胞功能的信号通路。在细胞核中，这些通路与已知的 膜隔室，但控制影响DNA修复，细胞存活和其他事件的应激反应。 激动剂激活PI 3 K信号传导通过IQGAP 1支架发生，该支架组装多个通路 包括PI 3-激酶和Erk途径。然而，IQGAP如何组装特定的信号通路并不清楚。 明白我们将专注于完整PI 3-激酶途径在IQGAP上的组装。这包括 将PI 4-激酶（PI 4KIII β）、I型PI 3 K PIP 5-激酶（PIPKI β）、PI 3 K、Ras、PDK 1和Akt组装成 脚手架值得注意的是，我们发现IQGAP 2和IQGAP 3也组装PI 3-激酶通路组件 但结果不同IQGAP 2是癌细胞中的肿瘤抑制因子，而IQGAP 1和IQGAP 3促进癌细胞的增殖。 PI 3 K信号传导和细胞增殖。在这里，我们将探讨受体如何刺激IQGAP的组装 信号通路，重点是EGF受体和IQGAP 1-PI 3-激酶和Erk通路。我们将 强调空间PI 3-激酶信号传导在内体区室在接近微管的联系， 微管相关蛋白4（MAP 4）与IQGAP 1和PI 3-激酶相互作用。之间的联系 胞质和核PI信号传导是PIPKI信号传导，其在胞质和核中产生PIP 2 核PI信号显着不与膜隔室。我们发现一个核武器 多聚腺苷酸聚合酶，Star-PAP（斑点靶向PIPKI调节的多聚腺苷酸聚合酶），与 PIPKI是磷脂酰肌醇-4，5-二磷酸（PIP 2）的底物，被激活。Star-PAP控制约40%的基因， 它受到许多信号的调节。最近，我们已经证明PIPKI β也与肿瘤抑制因子p53结合， p53是PIP 2效应子PIP 2的结合刺激p53与其他核因子的相互作用， 控制p53功能。Star-PAP和p53也受肌醇多磷酸多激酶（IPMK）调节。 产生与p53相关的磷脂酰肌醇-3，4，5-三磷酸（PIP 3），以及 使PIP 3去磷酸化。我们已经确定Star-PAP和p53是两个关键的核转录因子。 在应激信号传导期间的磷酸肌醇信号传导。本提案将重点讨论这一机制和影响， 应激途径对Star-PAP功能的影响。值得注意的是，PIPn与Star-PAP和p53紧密相关， 它对SDS-PAGE是稳定的，表明共价连接，我们将探索PIP 2如何连接到Star-PAP， 第53页。这种相互作用是共价的还是非常紧密的，能够抵抗变性？我们的发现指出了 对于胞质和核PI途径的潜在治疗控制，因为这些途径具有 在许多疾病过程中的基本影响，但重点是癌症。

PROJECT SUMMARY. This research seeks to understand spatial phosphoinositide signaling (PI) mechanisms in the cytosol and nucleus. These pathways have broad implications for cancer, neurodevelopmental disorders, diabetes, and several congenital diseases. In the cytosol, agonists, such as EGF (and many others), activate signaling pathways that control most cellular functions. In the nucleus, these pathways are separate from known membrane compartments but control stress responses that impact DNA repair, cell survival, and other events. Agonists activated PI3K signaling occurs through the IQGAP1 scaffold that assembles multiple pathways including the PI 3-kinase and Erk pathways. Yet, how the IQGAPs assemble specific signaling pathways is not understood. We will focus on the assembly of the full PI 3-kinase pathway on IQGAPs. This includes the assembly of the PI 4-kinase (PI4KIII), type I PIP 5-kinase (PIPKI), PI3K, Ras, PDK1 and Akt into the scaffold. Remarkably, we show that IQGAP2 and IQGAP3 also assemble the PI 3-kinase pathway components but with different outcomes. IQGAP2 is tumor suppressor in cancer cells whereas IQGAP1 and IQGAP3 promote PI3K signaling and cell proliferation. Here, we will explore how receptors stimulate the assembly of the IQGAPs signaling pathways with an emphasis on the EGF receptor and IQGAP1-PI 3-kinase and Erk pathways. We will emphasize spatial PI 3-kinase signaling at endosomal compartments at proximity to microtubules by linkage with microtubule associated protein 4 (MAP4) that interacts with IQGAP1 and PI 3-kinase. The link between the cytosolic and nuclear PI signaling is the PIPKI, which generates PIP2 in the cytosol and nucleus Nuclear PI signaling remarkably is not associated with membrane compartments. We showed that a nuclear poly(A) polymerase, Star-PAP (for speckle targeted PIPKI regulated-poly(A) polymerase), associates with PIPKI and is activated by phosphatidylinositol-4,5-bisphosphate (PIP2). Star-PAP controls ~40% of genes and is regulated by many signals. Recently, we have shown that PIPKI also binds to the tumor suppressor p53, and that p53 is a PIP2 effector. The binding of PIP2 stimulates p53’s interactions with other nuclear factors that control p53 function. Both Star-PAP and p53 are also regulated by inositol polyphosphate multikinase (IPMK) that generates phosphatidylinositol-3,4,5-trisphosphate (PIP3) associated with p53, and nuclear PTEN that dephosphorylates this PIP3. We have identified Star-PAP and p53 as two key effectors of nuclear phosphoinositide signaling during stress signaling. This proposal will focus on the mechanism and impact of this stress pathway on Star-PAP functions. Remarkably the PIPn is so tightly associated with Star-PAP and p53 that it is stable to SDS-PAGE suggesting a covalent linkage and we will explore how PIP2 is linked to Star-PAP and p53. Is this covalent or a very tight interaction that is resistant to denaturation? Our findings indicate new avenues for potential therapeutic control of both the cytosolic and nuclear PI pathways as these pathways have fundamental implications in many disease processes but with an emphasis on cancer.