A Novel Function for G6PD in Regulation of the Cancer Epigenome

G6PD 调节癌症表观基因组的新功能

• 批准号: 8772019
• 负责人: Maria Vogelauer
• 金额: $ 7.7万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-11 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8772019
• 关键词: Acetylation; Adenine Nucleotides; Affinity; Antineoplastic Agents; Antioxidants; Binding; Binding Sites; Biological; Biological Models; Biology; Breast Cancer Cell; Calcium; Calcium Channel; Cancer Biology; Cancer cell line; Cardiac; Cardiac Myocytes; Cardiomyopathies; Cell Nucleus; Cells; ChIP-seq; Chromatin; Coenzyme A; Communication; Coupled; Data; Data Analyses; Deacetylation; Development; Disease; Electron Transport; Elements; Enzymes; Epigenetic Process; Fatty Acids; Gene Expression; Gene Expression Regulation; Gene Structure; Generations; Genes; Genome; Genomics; Glucose-6-Phosphate; Glucosephosphate Dehydrogenase; HDAC1 gene; HDAC2 gene; Hepatocyte; Histone Acetylation; Histone Deacetylation; Histones; Homeostasis; In Vitro; Investigation; Kinetics; Laboratories; Lead; Link; Liver; Lysine; MDA MB 231; Malignant Neoplasms; Mammary Neoplasms; Mammary gland; Messenger RNA; Metabolic; Metabolic Pathway; Metabolism; Mus; NADH; NADP; Normal tissue morphology; Nuclear; Nucleic Acids; Nucleotides; Oxidation-Reduction; Pathway interactions; Pattern; Pentosephosphate Pathway; Pericardial effusion; Play; Probability; Process; Proteins; Reaction; Recurrence; Regulation; Reporting; Role; Testing; The Cancer Genome Atlas; Time; Transcriptional Regulation; abstracting; base; cancer cell; cell growth; deep sequencing; dehydroepiandrosterone; epigenome; genome-wide; human disease; in vivo; knock-down; macromolecule; malignant breast neoplasm; novel; public health relevance; research study; response; transcriptome sequencing; tumor progression

DESCRIPTION (provided by applicant): Glucose-6-phosphate dehydrogenase (G6PD) is the rate-limiting enzyme of the pentose phosphate pathway, the major metabolic pathway for generation of reducing equivalents for the cell. In an oxidation- reduction reaction, G6PD oxidizes glucose-6-phosphate and transfers the electrons to nicotinamide adenine dinucleotide phosphate (NADP+) to generate NADPH. NADPH provides reducing power for biosynthetic pathways that generate macromolecules such as synthesis of nucleic and fatty acids in the liver or the mammary glands. NADPH is also essential in maintaining antioxidant capacity of the cell. G6PD is normally considered to be a cytoplasmic enzyme where its activity is regulated by NADPH/NADP+ ratio. Increased G6PD activity has been reported for many diseases ranging from cardiomyopathy to cancers. We have recently discovered that NADPH, but not NADP+, NADH or NAD+, is an allosteric activator of class I histone deacetylases (HDACs) 1 and 2 in vitro. NADPH activates HDACs through a mixed activation kinetic, increasing the affinity of the HDAC enzyme for its histone substrate as well as the velocity of the deacetylation reaction. To provide in vivo support for our findings, we surmised that pharmacological inhibition of G6PD inside cells should lead to decreased NADPH levels and in turn to decreased HDAC activity and increased histone acetylation. Indeed, we observed that G6PD inhibition in MDA-MB-453 mammary cancer cells increases global histone acetylation by up to three fold. Another breast cancer cell lines, MDA-MB-231, did not show the same response to G6PD inhibition. Further investigation revealed that G6PD is highly expressed in MDA-MB-453 cells compared to MDA-MB-231 and that a detectable fraction of G6PD is localized in the nucleus with a further fraction strikingly bound to chromatin. In the 80 years since G6PD was discovered, only two reports indicate a potential presence of G6PD in the nucleus and none on chromatin. Moreover, our preliminary ChIP-seq analysis has revealed that G6PD binding to chromatin is not spurious but rather near genes with functions related to calcium homeostasis-an important pathway in breast cancer biology. In this application, we propose that G6PD binding to chromatin reveals an additional and significant function of G6PD that has not been characterized to any extent. We further postulate that G6PD may function at specific genomic loci to produce NADPH locally for HDAC activation and histone deacetylation. We now aim to determine the distribution of G6PD across the genome and its functional relevance to HDAC binding, histone acetylation and regulation of chromatin based processes such as gene expression. We believe our data will open up a new field of inquiry into the function of this critical metabolic enzyme and will uncover a remarkable juncture where metabolism and epigenetic regulation of the genome intersect.

描述（由申请人提供）：葡萄糖-6-磷酸脱氢酶（G6 PD）是戊糖磷酸途径的限速酶，戊糖磷酸途径是细胞产生还原当量的主要代谢途径。在氧化还原反应中，G6 PD氧化葡萄糖-6-磷酸并将电子转移到烟酰胺腺嘌呤二核苷酸磷酸（NADP+）以生成NADPH。NADPH为产生大分子的生物合成途径提供还原力，如肝脏或乳腺中核酸和脂肪酸的合成。NADPH在维持细胞的抗氧化能力方面也是至关重要的。G6 PD通常被认为是一种细胞质酶，其活性受NADPH/NADP+比率调节。已经报道了从心肌病到癌症的许多疾病的G6 PD活性增加。 我们最近发现，NADPH，而不是NADP+，NADH或NAD+，是I类组蛋白去乙酰化酶（HDACs）1和2的变构激活剂。NADPH通过混合活化动力学活化HDAC，增加HDAC酶对其组蛋白底物的亲和力以及脱乙酰化反应的速度。为了给我们的发现提供体内支持，我们推测细胞内G6 PD的药理学抑制应导致NADPH水平降低，进而导致HDAC活性降低和组蛋白乙酰化增加。事实上，我们观察到MDA-MB-453乳腺癌细胞中的G6 PD抑制使整体组蛋白乙酰化增加高达三倍。另一种乳腺癌细胞系MDA-MB-231对G6 PD抑制没有显示出相同的反应。进一步的研究表明，与MDA-MB-231相比，G6 PD在MDA-MB-453细胞中高度表达，并且G6 PD的可检测部分定位于细胞核中，另一部分与染色质显著结合。自G6 PD被发现以来的80年中，只有两份报告表明G6 PD可能存在于细胞核中，而染色质上没有。此外，我们初步的ChIP-seq分析表明，G6 PD与染色质的结合不是虚假的，而是与钙稳态相关功能的基因（乳腺癌生物学中的一个重要途径）。在本申请中，我们提出G6 PD与染色质的结合揭示了G6 PD的一个额外的和重要的功能，该功能尚未在任何程度上被表征。我们进一步推测，G6 PD可能在特定的基因组位点发挥作用，在HDAC激活和组蛋白去乙酰化中局部产生NADPH。我们现在的目标是确定G6 PD在整个基因组中的分布及其与HDAC结合，组蛋白乙酰化和基于染色质的过程如基因表达的调节的功能相关性。我们相信，我们的数据将为研究这种关键代谢酶的功能开辟一个新的领域， 这是基因组的新陈代谢和表观遗传调控相互交叉的一个重要时刻。