Komori K, Takagi Y, Sanada M, Lim T-H, Nakatsu Y, Tsuzuki T, Sekiguchi M, Hidaka M
Department of Molecular Biology, Biomolecular Engineering Research Institute, Suita, Japan.
Oncogene. 2009 Feb 26;28(8):1142-50. doi: 10.1038/onc.2008.462. Epub 2009 Jan 12.
O(6)-Methylguanine produced in DNA induces mutation due to its ambiguous base-pairing properties during DNA replication. To suppress such an outcome, organisms possess a mechanism to eliminate cells carrying O(6)-methylguanine by inducing apoptosis that requires the function of mismatch repair proteins. To identify other factors involved in this apoptotic process, we performed retrovirus-mediated gene-trap mutagenesis and isolated a mutant that acquired resistance to a simple alkylating agent, N-methyl-N-nitrosourea (MNU). However, it was still sensitive to methyl methanesulfonate, 1-(4-amino-2-methyl-5-pyrimidinyl)methyl-3-(2-chloroethyl)-3-nitrosourea, etoposide and ultraviolet irradiation. Moreover, the mutant exhibited an increased mutant frequency after exposure to MNU. The gene responsible was identified and designated Mapo1 (O(6)-methylguanine-induced apoptosis 1). When the expression of the gene was inhibited by small interfering RNA, MNU-induced apoptosis was significantly suppressed. In the Mapo1-defective mutant cells treated with MNU, the mitochondrial membrane depolarization and caspase-3 activation were severely suppressed, although phosphorylation of p53, CHK1 and histone H2AX was observed. The orthologs of the Mapo1 gene are present in various organisms from nematode to humans. Both mouse and human MAPO1 proteins expressed in cells localize in the cytoplasm. We therefore propose that MAPO1 may play a role in the signal-transduction pathway of apoptosis induced by O(6)-methylguanine-mispaired lesions.
DNA中产生的O(6)-甲基鸟嘌呤因其在DNA复制过程中模糊的碱基配对特性而诱发突变。为抑制这种结果，生物体拥有一种机制，通过诱导凋亡来清除携带O(6)-甲基鸟嘌呤的细胞，这一过程需要错配修复蛋白的功能。为鉴定参与该凋亡过程的其他因子，我们进行了逆转录病毒介导的基因捕获诱变，并分离出一个对简单烷基化剂N-甲基-N-亚硝基脲（MNU）具有抗性的突变体。然而，它对甲磺酸甲酯、1-(4-氨基-2-甲基-5-嘧啶基)甲基-3-(2-氯乙基)-3-亚硝基脲、依托泊苷和紫外线照射仍敏感。此外，该突变体在暴露于MNU后表现出增加的突变频率。鉴定出负责的基因并将其命名为Mapo1（O(6)-甲基鸟嘌呤诱导的凋亡1）。当该基因的表达被小干扰RNA抑制时，MNU诱导的凋亡被显著抑制。在用MNU处理的Mapo1缺陷突变体细胞中，尽管观察到p53、CHK1和组蛋白H2AX的磷酸化，但线粒体膜去极化和caspase-3激活被严重抑制。Mapo1基因的直系同源物存在于从线虫到人类的各种生物体中。在细胞中表达的小鼠和人类MAPO1蛋白均定位于细胞质。因此，我们提出MAPO1可能在由O(6)-甲基鸟嘌呤错配损伤诱导的凋亡信号转导途径中发挥作用。
