Metabolism and Epigenetics

项目摘要 该项目旨在揭示如何在基因编码中发现癌症驱动突变， 中枢代谢酶导致多种类型癌症的表观遗传后果， 这一领域的工作表明，影响这些酶之一的突变， 异柠檬酸脱氢酶，产生一种致癌代谢物2-羟基戊二酸（2-HG） 其可以抑制去除DNA和组蛋白的甲基修饰的酶。这 甲基化的靶点在癌症中发挥的关键作用尚不清楚。PI已经表明，同样的 人类癌症中发现的突变，当基因工程进入酵母酿酒酵母时， 产生2-HG。他还发现了一种酵母基因，该基因编码一种代谢2- Hg.使用一种巧妙的异染色质扰动分析， 通过这些突变似乎超稳定异染色质，而不是通过抑制DNA 去甲基化，但通过抑制赖氨酸36的特定甲基化物质的去甲基化， 组蛋白H3。拟议的实验通过直接的生物化学方法扩展了这些遗传推断。 试验.他们将测试2-HG是否影响染色质的任何其他结构， 建立了他们擅长的基因组学方法。进一步的目标扩大了他们的惊人 发现2-HG也由野生型细胞产生。通过利用系统的力量， 基因阵列（SGA）技术，他们将确定2-HG在细胞中的天然作用， 所有的基因，当突变时，使细胞对升高的2-HG敏感。酵母模型 将制备琥珀酸脱氢酶和琥珀酸脱氢酶的癌症驱动等位基因，并测试其活性。 表观遗传影响，如果发现，将建立影响的机制。基础上 成功与酵母模型的异柠檬酸脱氢酶突变体，PI已经完成了 成功的试验性筛选，到目前为止，通过以下途径鉴定了5个影响异染色质稳定性的基因： 对新陈代谢的影响，并建议完成估计恢复或50- 100个基因，代表了新陈代谢和 表观遗传学每个扰动的表观遗传效应背后的机制将被确定。

Project Summary This project is designed to reveal how the cancer-driver mutations found in the genes encoding central metabolic enzymes cause epigenetic consequences in the multiple types of cancers in which they are found. Work in the field indicates that mutations affecting one of these enzymes, isocitrate dehydrogenase, produce an onco-metabolite known a 2-hydroxyglutarate (2-HG) which can inhibit enzymes that remove methyl modifications of DNA and histones. Which methylated targets play critical roles in cancers is unclear. The PI has show that the very same mutations found in human cancer, when engineered into the yeast Saccharomyces cerevisiae produce 2-HG. He has also found a yeast gene that encodes an enzyme that metabolizes 2- HG. Using an ingenious assay for pertubations of heterochromatin, the overproduction caused by these mutations appears to hyper-stabilize heterochromatin, not by inhibiting DNA demethylation, but by inhibiting demethylation of a specific methylated species of lysine 36 on histone H3. The proposed experiments extend these genetic inferences by direct biochemical tests. They will test whether 2-HG is affecting any of the other structures of chromatin by well- established genomic methods that they are expert in. Further aims extend their surprising discovery that 2-HG is also made by wild-type cells. By harnessing the power of systematic genetic array (SGA) technology, they will identify the natural role of 2-HG in cells by identifying all genes that, when mutant, render cells sensitive to elevated 2-HG. Yeast models for the cancer-driving alleles of succinate dehydrogenase and fumarase will be made and tested for epigenetic impacts, and if found, the mechanism of the impact will be established. Building upon success with the yeast model of isocitrate dehydrogenase mutants, the PI has completed a successful pilot screen that identified 5 genes so far that affect heterochromatin stability through effects on metabolism, and propose completing the screen with an estimated recovery or 50- 100 genes, representing a near-comprehensive set of intersections betweeen metabolism and epigenetics. The mechanism behind each perturbations's epigenetic effect will be determined.