Cancer cell signaling through lipids complexed to proteins

描述(由申请者提供)：美国癌症协会2012年的报告显示，自1999年以来，美国大多数癌症的发病率正在下降或稳定。一些值得注意的例外是非洲裔美国人和西班牙裔美国人的肝癌，非裔美国女性的子宫内膜癌，50岁以下患者的结直肠癌，以及所有人口群体的胰腺癌。在所有这些组织中，NR5A核受体(NR5A1和NR5A2)介导的遗传程序是发育、分化和成年功能的基本决定因素。在分子水平上，这两个转录因子结合磷脂酰肌醇(PIP)，磷脂在PTEN依赖的癌症中对PI-3激酶信号转导是必不可少的。然而，关于这些脂质如何调节NR5A转录功能，或者这些机制的失调如何导致上述癌症，几乎一无所知。根据《科学》杂志目前正在修订的一篇手稿，我们假设与NR5As(和其他核蛋白)结合的PIP脂类直接由癌细胞中的PI3-激酶-肌醇多聚磷酸酶(IPMK)和PTEN脂磷酸酶重塑。这一假设明显背离了标准教条，即PI3-激酶和PTEN只作用于膜中的磷脂酰肌醇。在目标1中，我们将利用一个风险相对较低的项目，利用一个相对低风险的项目，利用一个相对低风险的项目，确定目前可用的小分子如何抑制IPMK，加速这些抑制剂的改进，并在结晶学、X射线衍射方法和分子结构确定方面培训PI。在目标2中，我们将使用突变分析来定位NR5A1和IPMK之间的界面，并尝试结晶NR5A/PIP/IPMK来确定这种复合体的界面结构。这种高风险的高回报结构被目标1对冲。在目标3中，我们将使用假设驱动的生物化学来鉴定作为PTEN和IPMK底物的新的蛋白质/PIP复合体。应聘者曾接受过蛋白质化学、酶学、生物化学和细胞生物学方面的广泛培训，但没有接受过任何结构生物学方面的培训。导师Holly Ingraham和顾问团队在结构生物学方面拥有丰富的经验，特别是在NR5A结构及其与胰腺癌的联系方面的Robert Fletterick，以及在激酶结构和膜癌生物学方面的Natalia Jura。候选人将利用K01奖项提供的培训，扩大他作为独立调查员的研究计划，为癌症研究开辟新的途径。

DESCRIPTION (provided by applicant): The American Cancer Society's 2012 report revealed that incidence rates are decreasing or stable for most cancers in the United States since 1999. Some notable exceptions are liver cancer in African Americans and Hispanics, endometrial cancer in African American women, colorectal cancer in patients under 50, and pancreatic cancer in every demographic. In all these tissues, NR5A nuclear receptors (NR5A1 and NR5A2) mediate genetic programs that are essential determinants of development, differentiation and adult function. At the molecular level, these two transcription factors bind phosphoinositides (PIPs), phospholipids that are essential to PI-3 kinase signaling in PTEN-dependent cancers. However almost nothing is known about how these lipids regulate NR5A transcriptional functions, or how dysregulation of those mechanisms contributes to the cancers mentioned above. Based on a manuscript currently in revision at Science, we hypothesize that PIP lipids bound to NR5As (and other nuclear proteins) are directly remodeled by the PI3- kinase inositol polyphosphate multikinase (IPMK) and the PTEN lipid phosphatase in cancer cells. This hypothesis is a clear departure from the standard dogma that PI3- kinases & PTEN only act on phosphoinositides in membranes. In Aim 1, we will determine how currently available small molecules inhibit IPMK using enzymology and crystallography, accelerating improvements of these inhibitors and training the PI in crystallography, X-ray diffraction methods and molecular structure determination, taking advantage of a relatively low-risk project. In Aim 2, we will use mutational analyses to map the interface between NR5A1 and IPMK, and attempt to crystallize NR5A/PIP/IPMK to determine the interfacial structure of this complex. This high-risk high-reward structure is hedged by Aim 1. In Aim 3, we will identify novel protein/PIP complexes that are substrates of PTEN and IPMK, using hypothesis-driven biochemistry. The candidate has had extensive training in protein chemistry, enzymology, biochemistry and cell biology, but has not had any training in structural biology. The mentor Holly Ingraham and advisory team have extensive experience in structural biology, particularly Robert Fletterick in NR5A structure and its links to pancreatic cancer, and Natalia Jura in kinase structure and membrane cancer biology. The candidate will expand his research program as an independent investigator with the training afforded by this K01 award, opening new avenues to cancer research.