叶酸循环和甜菜碱-同型半胱氨酸甲基转移酶都为小鼠囊胚中的DNA甲基化提供甲基基团。

Both the folate cycle and betaine-homocysteine methyltransferase contribute methyl groups for DNA methylation in mouse blastocysts.

作者信息

Zhang Baohua, Denomme Michelle M, White Carlee R, Leung Kit-Yi, Lee Martin B, Greene Nicholas D E, Mann Mellissa R W, Trasler Jacquetta M, Baltz Jay M

机构信息

*Ottawa Hospital Research Institute, Ottawa, Ontario, Canada; Departments of Obstetrics and Gynecology, and Cellular and Molecular Medicine, University of Ottawa Faculty of Medicine, Ottawa, Ontario, Canada; Department of Obstetrics and Gynecology, and Biochemistry, Schulich School of Medicine and Dentistry, Western University, London, Ontario, Canada; Children's Health Research Institute, London, Ontario, Canada; Developmental Biology and Cancer Program, University College London Institute of Child Health, London, United Kingdom; Research Institute of the McGill University Health Centre, Montréal Children's Hospital, Montréal, Quebec, Canada; and Departments of Human Genetics, Pediatrics, and Pharmacology and Therapeutics, McGill University, Montréal, Quebec, Canada.

出版信息

FASEB J. 2015 Mar;29(3):1069-79. doi: 10.1096/fj.14-261131. Epub 2014 Dec 2.

DOI:10.1096/fj.14-261131

PMID:25466894

Abstract

The embryonic pattern of global DNA methylation is first established in the inner cell mass (ICM) of the mouse blastocyst. The methyl donor S-adenosylmethionine (SAM) is produced in most cells through the folate cycle, but only a few cell types generate SAM from betaine (N,N,N-trimethylglycine) via betaine-homocysteine methyltransferase (BHMT), which is expressed in the mouse ICM. Here, mean ICM cell numbers decreased from 18-19 in controls to 11-13 when the folate cycle was inhibited by the antifolate methotrexate and to 12-14 when BHMT expression was knocked down by antisense morpholinos. Inhibiting both pathways, however, much more severely affected ICM development (7-8 cells). Total SAM levels in mouse blastocysts decreased significantly only when both pathways were inhibited (from 3.1 to 1.6 pmol/100 blastocysts). DNA methylation, detected as 5-methylcytosine (5-MeC) immunofluorescence in isolated ICMs, was minimally affected by inhibition of either pathway alone but decreased by at least 45-55% when both BHMT and the folate cycle were inhibited simultaneously. Effects on cell numbers and 5-MeC levels in the ICM were completely rescued by methionine (immediate SAM precursor) or SAM. Both the folate cycle and betaine/BHMT appear to contribute to a methyl pool required for normal ICM development and establishing initial embryonic DNA methylation.

摘要

全局DNA甲基化的胚胎模式首先在小鼠囊胚的内细胞团（ICM）中建立。甲基供体S-腺苷甲硫氨酸（SAM）在大多数细胞中通过叶酸循环产生，但只有少数细胞类型通过甜菜碱-同型半胱氨酸甲基转移酶（BHMT）从甜菜碱（N，N，N-三甲基甘氨酸）生成SAM，该酶在小鼠ICM中表达。在这里，当抗叶酸药物甲氨蝶呤抑制叶酸循环时，平均ICM细胞数从对照组的18 - 19个减少到11 - 13个，当通过反义吗啉代敲低BHMT表达时，减少到12 - 14个。然而，同时抑制这两条途径对ICM发育的影响要严重得多（7 - 8个细胞）。仅当两条途径都被抑制时，小鼠囊胚中的总SAM水平才显著降低（从3.1降至1.6 pmol/100个囊胚）。在分离的ICM中通过5-甲基胞嘧啶（5-MeC）免疫荧光检测到的DNA甲基化，单独抑制任何一条途径时受到的影响最小，但当同时抑制BHMT和叶酸循环时，DNA甲基化至少降低45 - 55%。甲硫氨酸（直接的SAM前体）或SAM完全挽救了对ICM中细胞数量和5-MeC水平的影响。叶酸循环和甜菜碱/BHMT似乎都对正常ICM发育和建立初始胚胎DNA甲基化所需的甲基库有贡献。