FOLATE DEFICIENCY AND ONE-CARBON METABOLISM

本提案的主要目标是确定是否存在缺陷， 叶酸，这是从头合成甲基所必需的， 导致类似的代谢结果， 组 饮食不含蛋氨酸和胆碱引起自发性 肝癌 蛋氨酸，一种干扰 甲基化，是一种致癌物质，但当注射到胆碱缺乏 在小鼠中，乙烯利迅速引起急性胰腺炎。 的一种解释 甲基缺乏的影响是S- 腺苷甲硫氨酸（SAM）转化为S-腺苷高半胱氨酸（SAH）。 当细胞内的甲基化反应被抑制时， SAM/SAH的比例降低，包括DNA的甲基化， 控制真核生物的基因表达。 这是由P.I. 叶酸缺乏导致SAM/SAH比率降低， 和胰腺，这是一样大的甲基基团所造成的 缺陷 这种SAM/SAH比率由甘氨酸N- 甲基转移酶（GNMT），其含有紧密结合的叶酸。 免疫组织化学研究表明，GNMT集中在 胰腺外分泌 高浓度的GNMT意味着甲基化 和叶酸对某些细胞类型特别重要。 这些 在具体目标方面进行了调查，这些目标是： 1.研究叶酸缺乏是否影响胰腺分泌 使用AR 42 J细胞系进行体内、原位和组织培养。 2.为了研究叶酸缺乏是否会影响 小分子（肌酸、磷脂）和大分子（DNA）。 3.为了将GNMT的免疫组织化学定位扩展到各种 组织。 4.目的：建立一种免疫组织化学方法， 组织中 叶酸的免疫组织化学定位将在 神经化学家用于脑定位的方法 神经传递素 快速叶酸缺乏症将在大鼠中产生， 喂养沃尔泽姆和克利福德的氨基酸限定饮食。 甲基化不足 将通过酶促掺入DNA的能力来评估 额外的甲基，通过改变poly A+ RNA的表达，和 通过改变DNA对限制性内切酶的反应。 叶酸缺乏时DNA甲基化不足会影响基因表达。 评估组织内特定细胞叶酸水平的能力 可以提供一种寻找局部叶酸缺乏的方法 吸烟者和口服避孕药者的肿瘤前病变。

The broad objective of this proposal is to determine whether a deficiency of folate, which is needed for the de novo synthesis of methyl groups, causes similar metabolic consequences as a deficiency of preformed methyl groups. Diets that contain no methionine and choline cause spontaneous liver cancer. Ethionine, an analogue of methionine that interferes with methylation, is a carcinogen, but when injected to choline deficient mice, ethionine rapidly causes acute pancreatitis. One explanation for the effects of methyl deficiency has been the decreased ratio of S- adenosylmethionine (SAM) to S-adenosylhomocysteine (SAH) that results. A wide variety of cellular methylation reactions are inhibited when the ratio of SAM/SAH is decreased, including the methylation of DNA which controls gene expression in eukaryotes. It has been shown by the P.I. that folate deficiency produces a decrease in the SAM/SAH ratio, in liver and pancreas, which is as great as that caused by methyl group deficiency. This SAM/SAH ratio is regulated by the enzyme, glycine N- methyltransferase (GNMT), which contains tightly bound folate. Immunohistochemical studies have shown that GNMT is concentrated in the exocrine pancreas. A high concentration of GNMT implies that methylation and folate are particularly important for certain cell types. These aspects are investigated in the specific aims which are: 1. To investigate whether folate deficiency affects pancreatic secretion in vivo, in situ and in tissue culture using the AR42J cell line. 2. To investigate whether folate deficiency affects the methylation of small molecules (creatine, phospholipids) and macromolecules (DNA). 3. To extend the immunohistochemical localization of GNMT to a variety of tissues. 4. To develop an immunohistochemical method for detecting folate within tissues. Immunohistochemical localization of folate will be modeled after the method used by the neurochemists for the localization of brain neurotransmitters. Rapid folate deficiency will be produced in rats by feeding the amino defined diet of Walzem and Clifford. Undermethylation of DNA will be evaluated by the ability to enzymatically incorporate additional methyl groups, by an altered expression of poly A+ RNA, and by an altered response of DNA to restriction endonucleases. Undermethylation of DNA in folate deficiency can affect gene expression. The ability to estimate folate levels of specific cells within tissues may provide a means of looking for localized folate deficiency as a cause of pre-neoplastic lesions in smokers and oral-contraceptive users.