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: 8910675
• 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/8910675
• 关键词: Acetylation; Adenine Nucleotides; Affinity; Antineoplastic Agents; Antioxidants; Binding; Binding Sites; Biological; Biological Models; Biology; Breast Cancer Cell; Breast Cancer cell line; Calcium; Calcium Channel; Cancer Biology; Cardiac; Cardiac Myocytes; Cardiomyopathies; Cell Nucleus; Cells; ChIP-seq; Chromatin; Coenzyme A; Communication; Coupled; Data; Data Analyses; Deacetylation; Development; Disease; Electron Transport; Elements; Enzymes; 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; Metastatic breast cancer; 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; epigenetic regulation; 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（G6PD）是戊糖磷酸途径的限速酶，戊糖磷酸途径是细胞产生还原性当量的主要代谢途径。在氧化还原反应中，G6PD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+）上生成NADPH。NADPH为产生大分子的生物合成途径提供还原能力，如肝脏或乳腺中核酸和脂肪酸的合成。NADPH对于维持细胞的抗氧化能力也是必不可少的。G6PD通常被认为是一种细胞质酶，其活性受NADPH/NADP+比值调节。从心肌病到癌症，许多疾病都报道了G6PD活性增高。我们最近在体外发现NADPH是I类组蛋白去乙酰化酶（HDACs） 1和2的变构激活剂，而不是NADP+、NADH或NAD+。NADPH通过混合激活动力学激活HDAC，增加HDAC酶对其组蛋白底物的亲和力以及去乙酰化反应的速度。为了为我们的研究结果提供体内支持，我们推测细胞内G6PD的药理抑制应导致NADPH水平降低，进而降低HDAC活性和增加组蛋白乙酰化。事实上，我们观察到G6PD在MDA-MB-453乳腺癌细胞中的抑制使全局组蛋白乙酰化增加了三倍。另一种乳腺癌细胞系MDA-MB-231对G6PD抑制没有表现出相同的反应。进一步的研究表明，与MDA-MB-231相比，G6PD在MDA-MB-453细胞中高表达，并且G6PD的可检测部分定位于细胞核中，另一部分明显与染色质结合。在G6PD被发现的80年里，只有两篇报道指出G6PD可能存在于细胞核中，而没有报道指出在染色质上存在G6PD。此外，我们初步的ChIP-seq分析显示，G6PD与染色质的结合并不是假的，而是靠近与钙稳态相关的基因，钙稳态是乳腺癌生物学的一个重要途径。在这个应用中，我们提出G6PD与染色质的结合揭示了G6PD的一个额外的和重要的功能，这个功能在任何程度上都没有被表征。我们进一步假设G6PD可能在特定的基因组位点上起作用，在局部产生NADPH以激活HDAC和组蛋白去乙酰化。我们现在的目标是确定G6PD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）的调控的功能相关性。我们相信，我们的数据将为这种关键代谢酶的功能开辟一个新的研究领域，并将揭示