Alkylation-induced apoptosis of embryonic stem cells in which the gene for DNA-repair, methyltransferase, had been disrupted by gene targeting.
Alkylation-induced apoptosis of embryonic stem cells in which the gene for DNA-repair, methyltransferase, had been disrupted by gene targeting.
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烷基化诱导胚胎干细胞凋亡，其中 DNA 修复基因甲基转移酶已被基因靶向破坏。
DOI：
10.1093/carcin/18.5.889
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发表时间：
1997
期刊：
影响因子：
4.7
通讯作者：
M. Sekiguchi
中科院分区：
文献类型：
--
作者：
Y. Tominaga;T. Tsuzuki;A. Shiraishi;H. Kawate;M. Sekiguchi
o6 -甲基鸟嘌呤-DNA甲基转移酶（MGMT）催化甲基从o6 -甲基鸟嘌呤和o4 -甲基胸腺嘧啶的烷基化DNA转移到自己的分子中，从而在单步反应中修复致突变前的损伤。利用基因靶向技术，我们培育了缺乏甲基转移酶的小鼠胚胎干细胞系。定量免疫印迹分析和酶分析显示，两个等位基因都被破坏的MGMT-/-细胞不含甲基转移酶蛋白，而一个完整等位基因（MGMT+/-）的细胞所含的甲基转移酶蛋白约为亲本MGMT+/+细胞的一半。MGMT-/-细胞对简单的烷基化剂n -甲基-n '-硝基-n -亚硝基胍（MNNG）和n -甲基-n -亚硝基脲（MNU）具有极高的敏感性，而MGMT+/-细胞对这些剂的敏感性略高于正常细胞。暴露于相对低剂量的MNNG后，MGMT-/-细胞发生高频率的突变。dna的电泳分析和细胞的荧光染色显示，MNNG处理的MGMT-/-细胞发生凋亡死亡，这发生在细胞增殖第二周期G2-M阻滞后。
An enzyme O6-methylguanine-DNA methyltransferase (MGMT) catalyzes transfer of a methyl group from O6-methylguanine and O4-methylthymine of alkylated DNA to its own molecule, thereby repairing the pre-mutagenic lesions in a single step reaction. Making use of gene targeting, we developed mouse embryonic stem (ES) cell lines deficient in the methyltransferase. Quantitative immunoblot analysis and enzyme assay revealed that MGMT-/- cells, in which both alleles were disrupted, contained no methyltransferase protein while cells with one intact allele (MGMT+/-) contained about half the amount of protein carried by the parental MGMT+/+ cells. MGMT-/- cells have an extremely high degree of sensitivity to simple alkylating agents, N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and N-methyl-N-nitrosourea (MNU), whereas MGMT+/- cells are slightly more sensitive to these agents, as compared with findings from normal cells. A high frequency of mutation was induced in MGMT-/- cells on exposure to a relatively low dose of MNNG. Electrophoretic analyses of the DNAs as well as fluorochrome staining of the cells revealed that MGMT-/- cells treated with MNNG undergo apoptotic death, which occurs after G2-M arrest in the second cycle of cell proliferation.
DOI：
10.1073/pnas.90.14.6424
发表时间：
1993-07-15
影响因子：
11.1
作者：
KAT, A;THILLY, WG;MODRICH, P
通讯作者：
MODRICH, P
人淋巴母细胞株暴露于 N-甲基-N-硝基-N-亚硝基胍后的细胞周期扰动和细胞死亡。
DOI：
--
发表时间：
1989
期刊：
The American journal of pathology
影响因子：
--
作者：
Black,KA;McFarland,RD;Grisham,JW;Smith,GJ
通讯作者：
Smith,GJ
影响因子：
14.9
作者：
HARRIS, LC;POTTER, PM;BRENT, TP
通讯作者：
BRENT, TP