Biochemistry of the lysine beta-hydroxybutyrylation pathway

新出现的证据表明，肥胖、能量代谢、营养和表观遗传之间存在密切联系。 机制表观遗传学变化，如动态组蛋白修饰，与细胞增殖有关。 代谢和糖尿病并发症。然而，介导串扰的分子机制 代谢和表观遗传学之间的联系仍然不完全清楚。我们最近发现， 全面验证了一种新的、进化上保守的赖氨酸修饰，赖氨酸β（β）- 核心组蛋白上的羟基丁酰化（Kbhb）。我们在组蛋白上检测到44个非冗余Kbhb标记， 鉴定Sirt2酶为第一种去除组蛋白Kbhb的酶。Kbhb的水平非常动态， 受生理条件的影响（例如，饥饿和I型糖尿病）和营养来源。增加 β-羟基丁酸酯（也称为3-羟基丁酸酯）水平导致组蛋白Kbhb增加，可能是通过 β-羟基丁酸转化为β-羟基丁酰辅酶A。有趣的是，组蛋白Kbhb富含活性 基因启动子，以及在长期饥饿期间发生的H3K9bhb水平增加与 基因在饥饿响应代谢途径中上调，因此代表了一种新的表观遗传学 将新陈代谢与基因表达结合的调节标记。β-羟基丁酸酯是 “酮体”，它已被用于几十个抗癌临床试验，作为一种潜在的治疗， 与其他代理商的组合。β-羟基丁酸酯的血浆/细胞浓度可增加至20 在饥饿和病理条件下，如糖尿病（DM）和酒精性肝损伤 这可以驱动组蛋白Kbhb的形成。已知高酮血症和酮症酸中毒会增加以下风险： 患者的发病率和死亡率。因此，Kbhb途径的分子表征不仅将改善我们的研究， 了解表观遗传机制，但也表征β-羟基丁酸在 生理病理学我们假设Kbhb途径在分子上不同于赖氨酸乙酰化 通路因此，我们建议通过定义其关键调控元件来表征Kbhb途径， 包括其底物，一套独特的调节酶和直接结合蛋白，从而奠定了 为研究其生物学功能奠定了基础。在这项研究中，我们将采用一种综合战略， 酶学、化学生物学、生物化学和蛋白质组学方法。我们的团队，Zhao实验室和 科尔实验室，是很好的定位，以执行这一项目，因为我们的综合专业知识，在这些 以及我们已经获得的有关初步数据。在这份提案中，我们将首先 使用定量分析方法全面识别和量化含KbHb底物的动态变化， 蛋白质组学方法。然后，我们将确定和表征KbHb调节酶，可以添加或删除 Kbhb。我们将最终确定和确认组蛋白Kbhb肽的直接蛋白结合剂。

Emerging lines of evidence suggest a close link between obesity, energy metabolism, nutrients and epigenetic mechanism. Epigenetic changes, such as dynamic histone modifications, are associated with cellular metabolism and diabetic complications. Nevertheless, the molecular mechanisms mediating the crosstalk between metabolism and epigenetics remain incompletely understood. We recently discovered and comprehensively validated a new, evolutionarily-conserved lysine modification, lysine beta(β)- hydroxybutyrylation (Kbhb), on core histones. We detected 44 non-redundant Kbhb marks on histones and identified Sirt2 enzyme as the first enzyme to remove histone Kbhb. Levels of Kbhb are very dynamic and are influenced by physiological conditions (e.g., starvation and type I diabetes) and nutrition sources. Increased levels of β-hydroxybutyrate (also called 3-hydroxybutyrate) lead to increased histone Kbhb, presumably via conversion of β-hydroxybutyrate to β-hydroxybutyryl CoA. Interestingly, histone Kbhb is enriched in active gene promoters, and the increased H3K9bhb levels that occur during prolonged starvation are associated with genes up-regulated in starvation-responsive metabolic pathways, thus representing a new epigenetic regulatory mark that couples metabolism to gene expression. β-Hydroxybutyrate is a key component of “ketone bodies” and it has been employed in dozens of anti-cancer clinical trials as a potential therapeutic in combination with other agents. The plasma/cellular concentration of β-hydroxybutyrate can increase up to 20 mM during starvation and in pathological conditions such as diabetes mellitus (DM) and alcoholic liver damage and this can drive histone Kbhb formation. Hyperketonemia and ketoacidosis are known to increase the risk of morbidity and mortality in patients. Thus, molecular characterization of Kbhb pathway will not only improve our understanding of epigenetic mechanism but also characterize functions of β-hydroxybutyrate in physiopathology. We hypothesize that the Kbhb pathway is molecularly distinct from the lysine acetylation pathway. We therefore propose to characterize the Kbhb pathway by defining its key regulatory elements, including its substrates, a unique set of regulatory enzymes and direct binding proteins, thus laying a foundation for studying its biology functions. We will use an integrated strategy in this study involving enzymology, chemical biology, biochemistry and proteomics approaches. Our team, the Zhao laboratory and the Cole laboratory, is well positioned to carry out this project, because of our combined expertise in these areas and the relevant preliminary data that we have already obtained. In this proposal, we will first comprehensively identify and quantify dynamic changes of Kbhb-containing substrates using a quantitative proteomics approach. We will then identify and characterize Kbhb-regulatory enzymes that can add or remove Kbhb. We will finally identify and confirm the direct protein binders of histone Kbhb peptides.