Phosphoinositide Signaling To and Within the Nucleus

进入细胞核和细胞核内的磷酸肌醇信号传导

• 批准号: 8059297
• 负责人: Richard A. Anderson
• 金额: $ 10万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-15 至 2011-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8059297
• 关键词: 1,2-diacylglycerol; 1-Phosphatidylinositol 3-Kinase; Amino Acid Sequence; Apolipoprotein E; Biological; Biological Assay; Cardiovascular Diseases; Cell Nucleus; Cell physiology; Cells; Chromatin; Code; Complex; Cytoplasmic Granules; DNA Polymerase II; Data; Diglycerides; Elements; Enzymes; Eukaryotic Cell; Gene Expression; Gene Proteins; Gene Targeting; Generations; Genes; Goals; Head; Health; Holoenzymes; Human; Hydrolysis; Hydroxyl Radical; Immunoprecipitation; In Vitro; Inositol; Lipids; Mass Spectrum Analysis; Membrane; Messenger RNA; Metabolism; Microarray Analysis; NAD(P)H dehydrogenase (quinone) 1, human; NQO1 gene; Names; Nerve Degeneration; Nuclear; Nuclear Protein; Nuclear Proteins; Organism; Oxidative Stress; Oxidative Stress Pathway; Pathway interactions; Phosphatidylinositol 4,5-Diphosphate; Phosphatidylinositol Phosphates; Phosphatidylinositols; Phosphoric Monoester Hydrolases; Phosphotransferases; Play; Poly A; Poly(A) Tail; Polyadenylation; Polyadenylation Pathway; Polynucleotide Adenylyltransferase; Positioning Attribute; Process; Production; Protein Isoforms; Proteins; Pulmonology; RNA; RNA 3' End Processing; RNA Sequences; Regulation; Role; Screening procedure; Second Messenger Systems; Signal Pathway; Signal Transduction; Signal Transduction Pathway; Site; Small Interfering RNA; Specificity; Stream; Stroke; Structure; Substrate Specificity; Tail; Testing; Transcript; Transduction Gene; Transplantation; Work; Yeasts; biological adaptation to stress; cancer therapy; crosslink; heme oxygenase-1; human disease; in vivo; interest; mRNA Expression; mRNA Precursor; member; novel; phosphatidylinositol 5-phosphate; polyadenosine; polyadenylated messenger RNA; protein protein interaction; public health relevance; response; second messenger; yeast two hybrid system

DESCRIPTION (provided by applicant): Phosphoinositide signaling regulates all eukaryotic cells. In the phosphoinositide cycle, phosphatidylinositol (PI) is sequentially phosphorylated on the fourth and fifth hydroxyl of the myo-inositol ring by PI kinases and then phosphatidylinositol-phosphate kinases (PIPKs), forming phosphatidylinositol-4,5-bisphosphate (PIP2). PIP2 is a direct lipid messenger for many cellular responses and is an essential precursor to many other phosphatidylinositol-derived second messengers. Remarkably phosphoinositide signaling occurs within the nucleus where PIP2 is spatially generated at structures called nuclear speckles. Speckles have no identified membrane, but contain proteins and enzymes with roles in mRNA processing. PIPKs signal by interacting with effectors of the PIPn that they generate. PIPKI1 is an isoform that makes PIP2 that is present in the nucleus at speckles. PIP2 is also generated at speckles and may regulate activities of mRNA processing enzymes. In this context, we have discovered that PIPKI1 interacts with a novel poly A polymerase (PAP) now called Star-PAP. Star-PAP is dramatically and specifically stimulated by PIP2. Star-PAP and PIPKI1 are regulated by oxidative stress response and this regulates the expression of stress response mRNAs, including heme oxygenase-1 (HO-1), apolipoprotein E (APOE) and NAD(P)H:quinone oxidoreductase (NQO1). We hypothesize that PIPKI1 and Star-PAP function as a polyadenylation complex that associates with the transcriptional machinery required for 3'-processing of pre-mRNA transcripts. This novel polyadenylation complex is uniquely regulated by phosphoinositide signals and is required in vivo for 3'- processing of select mRNA transcripts, resulting in a novel mechanism to regulate gene expression. We will test this hypothesis with the following focused specific aims: 1. Study enzymatic activity of Star- PAP and regulation by phosphoinositides. Functional domains in Star-PAP that modulate specificity toward RNA substrates will be revealed. 2. Characterize Star-PAP complex assembly down stream of oxidative stress signals. The role of functional domains in Star-PAP will be defined. 3. The mechanisms for Star-PAP regulation of mRNAs in vivo will be investigated with an emphasis on the interaction with genes and mRNAs. It will be determined if the poly(A) tail generated by Star-PAP different than that by canonical PAP. The regulation of expression of the stress response proteins HO-1, APOE, and NQO1 play key biological roles that have dramatic implications for many aspects of human health including cardiovascular disease, transplantation, neurodegeneration and stroke, cancer therapy, and pulmonary medicine. The regulation of pre-mRNA polyadenylation by phosphoinositide signals via PIPKI1 and Star-PAP is an incredibly novel finding that has many implications for nuclear signal transduction and gene expression. Public Health Relevance: The expression of cellular proteins from genes occurs through messenger RNAs (mRNAs) made by each gene and this requires that the mRNA have a polyadenosine tail. This tail is required before the mRNA can make cellular proteins. We have discovered a new enzyme that uniquely makes these tails and this enzyme works to generate mRNAs and the encoded proteins. Most interesting and paradigm shifting is the fact that this process is regulated by a lipid messenger called phosphatidylinositol-4,5-bisphosphate. The genes whose expression this novel pathway controls are heme oxygenase-1 (HO-1), apolipoprotein E (APOE) and NAD(P)H:quinone oxidoreductase (NQO1). The control of these genes has dramatic implications for many aspects of human health including cardiovascular disease, transplantation, neurodegeneration, cancer therapy, and pulmonary medicine.

描述（由申请人提供）：磷酸肌醇信号传导调节所有真核细胞。在磷酸肌醇周期中，PI激酶和磷脂性激酶的第四和第五羟基磷酸化磷酸化（PI）在肌醇环的第四和第五羟基上被磷酸化，然后在磷脂酰氨基辛醇 - 磷酸磷酸激酶（PIPKS）上磷酸化，形成磷脂型磷脂型磷脂型磷脂型磷脂型4,4,4,4,5-磷酸盐。 PIP2是许多细胞反应的直接脂质使者，也是许多其他磷脂酰肌醇衍生的第二使者的重要先驱。明显的磷酸肌醇信号传导发生在核中，其中PIP2在称为核斑点的结构上是空间产生的。斑点没有鉴定的膜，但含有在mRNA加工中具有作用的蛋白质和酶。 PIPK信号通过与它们产生的PIPN的效应子相互作用。 PIPKI1是一种使斑点核中存在的PIP2的同工型。 PIP2还在斑点上产生，可能调节mRNA加工酶的活性。在这种情况下，我们发现PIPKI1与现在称为Star-PAP的新型聚合物酶（PAP）相互作用。 Star-PAP由PIP2急剧，特异性刺激。 Star-PAP和PIPKI1受氧化应激反应的调节，这调节了应力反应mRNA的表达，包括血红素加氧酶-1（HO-1），载脂蛋白E（APOE）E（APOE）和NAD（P）H：奎因酮氧化还原酶（NQO1）。我们假设PIPKI1和Star-PAP作为一种多腺苷酸化络合物，该复合物与前MRNA转录物进行3'SCORACESS所需的转录机械相关。这种新型的聚腺苷酸化复合物受磷酸肌醇信号的独特调节，在体内需要进行3'-精选mRNA转录本的处理，从而产生了一种新的机制来调节基因表达。我们将以以下针对特定的目的检验这一假设：1。研究星木和调节磷酸肌醇的酶促活性。 Star-PAP中的功能域将显示对RNA底物调节特异性的特异性。 2。表征星-PAP复合物组件下方的氧化应激信号流。将定义功能域在星-PAP中的作用。 3。将研究体内mRNA的星-PAP调节的机制，重点是与基因和mRNA的相互作用。将确定由星-A产生的聚（A）尾巴与规范PAP产生的尾巴不同。压力反应蛋白HO-1，APOE和NQO1的表达的调节扮演着关键的生物学作用，对人类健康的许多方面具有巨大影响，包括心血管疾病，移植，神经变性和中风，癌症治疗，癌症治疗和肺部医学。通过PIPKI1和Star-PAP通过磷酸肌醇信号对MRNA聚腺苷酸化的调节是一个令人难以置信的新颖发现，对核信号转导和基因表达有许多影响。公共卫生相关性：来自基因的细胞蛋白的表达是通过每个基因制成的信使RNA（mRNA）发生的，这要求mRNA具有多腺苷尾巴。在mRNA可以制成细胞蛋白之前，需要此尾巴。我们发现了一种新的酶，该酶独特地制造了这些尾巴，该酶可以生成mRNA和编码的蛋白质。最有趣和最范式的转移是，该过程由称为磷脂酰肌醇-4,5-双磷酸盐的脂质信使调节。这种新型途径控制的基因是血红素氧酶-1（HO-1），载脂蛋白E（APOE）和NAD（P）H：喹酮氧化还原酶（NQO1）。这些基因的控制对人类健康的许多方面都具有巨大的影响，包括心血管疾病，移植，神经退行性，癌症治疗和肺医学。