Folate and S-adenosyimethionine in Methyl Group and One-carbon Metabolism

描述（由申请人提供）：甲基化的调节与叶酸代谢密切相关。这两个代谢过程通过甘氨酸N-甲基转移酶连接，该酶在肝脏、胰腺外分泌和前列腺中含量丰富，在肾脏和肠粘膜中含量较低。它通过S-腺苷甲硫氨酸（NAMet）对甘氨酸进行甲基化，形成N-甲基甘氨酸和S-腺苷高半胱氨酸（NAHcy）。该酶的功能被认为是调节β-Met与β-Hcy的比率。α Met是甲基的主要载体，α Met/α Hcy比值是细胞甲基化能力的指标。该酶还在体内和体外紧密结合5-甲基-四氢叶酸五谷氨酸，其作为酶的抑制剂。这将预先形成的甲基（如甲硫氨酸）的可用性与通过叶酸介导的反应从头合成甲基联系起来。我们开发了一种甘氨酸N-甲基转移酶敲除小鼠，与WT小鼠相比，其肝脏和血浆中具有非常高水平的S-腺苷甲硫氨酸。初步数据显示，肝脏DNA并非整体高度甲基化。这些-/-小鼠自发地发展脂肪肝。我们的第一个假设是，来自具有高水平甘氨酸N-甲基转移酶的-/-动物的肝外组织也将具有异常的组织学和升高的甘氨酸N-甲基转移酶。我们的第一个具体目标是检查组织学并测量WT和-/-小鼠组织中的甲硫氨酸代谢物。我们的第二个假说是基因的启动子区域将被高甲基化，影响关键蛋白质的表观遗传表达。我们的第二个具体目标是测量WT和小鼠组织中参与脂肪性肝炎和甲硫氨酸代谢发展的基因的DNA的整体甲基化和启动子区域。甘氨酸N-甲基转移酶在外分泌组织中含量最丰富。我们的第三个假设是甘氨酸N-甲基转移酶在外分泌中很重要，我们的第三个具体目标是测量WT和-/-小鼠的外分泌。肝脏中最丰富的叶酸辅酶是5-甲基四氢叶酸。肝脏中的大多数叶酸与甘氨酸N-甲基转移酶紧密结合。我们的第四个假设是，在缺乏这种蛋白质的情况下，不同形式的叶酸将被重新分配，并导致叶酸介导的反应发生改变。我们的第四个具体目标是测量WT和-/-小鼠的肝脏、胰腺和前列腺中各种形式的叶酸的分布，以及叶酸缺乏时是否发生变化。在-/-动物中，HDMet的水平非常高，表明蛋白质可能过度甲基化。我们的第五个假设是，在-/-动物中，他的-音调可能是高甲基化的。我们的第五个具体目标是检查WT和-/-小鼠中组蛋白的甲基化。我们的第六个假设是，有可能通过给予作为甲基受体的化合物来降低组织中的高水平的蛋氨酸，并降低-/-小鼠肝脏疾病的严重程度。我们的第六个具体目标是确定是否给予烟酰胺或胍乙酸将做到这一点。公共卫生相关性：该提案“叶酸和S-腺苷甲硫氨酸在甲基组和一碳代谢”处理的损失的影响的酶，甘氨酸N-甲基转移酶，一个主要的酶在肝脏，胰腺和前列腺。我们在几个儿童中发现了这种酶的突变，导致中度肝损伤，并开发了一种完全缺乏甘氨酸N-甲基转移酶的小鼠模型，随着年龄的增长，这种酶会发展为严重的肝脏疾病。该提案的目标之一是评估几种常见天然化合物在该小鼠模型中减少肝损伤发展的能力，从而提高治疗患有缺陷型甘氨酸N-甲基转移酶的人并预防严重肝损伤的可能性。

DESCRIPTION (provided by applicant): Regulation of methylation is intimately associated with folate metabolism. These two metabolic processes are connected by the enzyme, glycine N-methyltransferase, which is abundant in liver, exocrine pancreas and prostate as well as in kidney and intestinal mucosa in lower amounts. It carries out the methylation of glycine by S-adenosylmethionine (AdoMet) to form N-methylglycine and S-adenosylhomocysteine (AdoHcy). The function of the enzyme is believed to be regulation of the ratio of AdoMet to AdoHcy. AdoMet is the major do- nor of methyl groups and the AdoMet/AdoHcy ratio an index of the methylating capacity of the cell. The enzyme also tightly binds 5-methyl-tetrahydrofolate pentaglutamate in vivo and in vitro, which acts as an inhibitor of the enzyme. This links the availability of preformed methyl groups, as methionine, to the de novo synthesis of methyl groups via folate-mediated reactions. We developed a glycine N-methyltransferase knockout mouse that has very high levels of S-adenosylmethionine in the livers and plasma compared to WT mice. Preliminary data showed that liver DNA is not globally hypermethylated. These -/- mice spontaneously develop fatty liver. Our First Hypothesis is that extra-hepatic tissues from -/- animals with high levels of glycine N- methyltransferase will also have abnormal histology and elevated AdoMet. Our First Specific Aim is to ex- amine histology and measure methionine metabolites in tissues from WT, and -/- mice. Our Second Hy- pothesis is that promoter regions of genes will be hypermethylated affecting the epigenetic expression of key proteins. Our second Specific Aim is to measure DNA for global methylation and the promoter regions of genes involved in the development of steatohepatitis and methionine metabolism in tissue from WT and mice. Glycine N-methyltransferase is most abundant in exocrine tissue. Our Third Hypothesis is that glycine N-methyltransferase is important in exocrine secretion and our Third Specific Aim is to measure exocrine se- cretion in WT and -/- mice. The most abundant folate coenzyme in liver is 5-methyltetrahydrofolate. Most of the folate in the liver is tightly bound to glycine N-methyltransferase. Our Fourth Hypothesis is that in the absence of this protein the different forms of folate will be redistributed and result in alterations in folate-mediated reac- tions. Our Fourth Specific Aim is to measure the distribution of the various forms of folate in liver, pancreas and prostate of WT and -/- mice and also whether it is changed in folate deficiency. The levels of AdoMet are very high in -/- animals suggesting that proteins may be overmethylated. Our Fifth Hypothesis is that his- tones may be hypermethylated in -/- animals. Our Fifth Specific Aim is to examine the methylation of histones in WT and -/- mice. Our Sixth Hypothesis is that it is possible to reduce the high levels of AdoMet in tissues by administration of compounds that serve as acceptors of methyl groups and reduce the severity of liver dis- ease in -/- mice. Our Sixth Specific Aim is to determine whether administration of nicotinamide or guanidi- noacetate will do this. PUBLIC HEALTH RELEVANCE: This proposal "Folate and S-adenosylmethionine in Methyl Group and One-carbon Metabolism" deals with the effects of the loss of the enzyme, glycine N-methyltransferase, a major enzyme in liver, pancreas and the prostate gland. We have discovered mutations of this enzyme in several children that result in moderate liver damage and have developed a mouse model that has a complete absence of glycine N-methyltransferase that progresses to severe liver disease as they get older. One of the goals of this proposal is to evaluate the use of several common natural compounds for their ability to diminish the development of liver damage in this mouse model thus raising the possibility of treating humans with defective glycine N-methyltransferase and prevent serious liver damage.