Bae J-B, Mukhopadhyay S S, Liu L, Zhang N, Tan J, Akhter S, Liu X, Shen X, Li L, Legerski R J
Department of Cancer Genetics, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA.
Oncogene. 2008 Aug 28;27(37):5045-56. doi: 10.1038/onc.2008.139. Epub 2008 May 12.
The removal of DNA interstrand cross-links (ICLs) has proven to be notoriously complicated due to the involvement of multiple pathways of DNA repair, which include the Fanconi anemia/BRCA pathway, homologous recombination and components of the nucleotide excision and mismatch repair pathways. Members of the SNM1 gene family have also been shown to have a role in mediating cellular resistance to ICLs, although their precise function has remained elusive. Here, we show that knockdown of Snm1B/Apollo in human cells results in hypersensitivity to mitomycin C (MMC), but not to IR. We also show that Snm1B-deficient cells exhibit a defective S phase checkpoint in response to MMC, but not to IR, and this finding may account for the specific sensitivity to the cross-linking drug. Interestingly, although previous studies have largely implicated ATR as the major kinase activated in response to ICLs, we show that it is activation of the ATM-mediated checkpoint that is defective in Snm1B-deficient cells. The requirement for Snm1B in ATM checkpoint activation specifically after ICL damage is correlated with its role in promoting double-strand break formation, and thus replication fork collapse. Consistent with this result Snm1B was found to interact directly with Mus81-Eme1, an endonuclease previously implicated in fork collapse. In addition, we also show that Snm1B interacts with the Mre11-Rad50-Nbs1 (MRN) complex and with FancD2 further substantiating its role as a checkpoint/DNA repair protein.
由于DNA修复的多种途径参与其中，包括范可尼贫血/BRCA途径、同源重组以及核苷酸切除和错配修复途径的组成部分，DNA链间交联（ICL）的去除已被证明极其复杂。SNM1基因家族的成员也已被证明在介导细胞对ICL的抗性中发挥作用，尽管它们的确切功能仍然难以捉摸。在这里，我们表明在人类细胞中敲低Snm1B/Apollo会导致对丝裂霉素C（MMC）敏感，但对电离辐射（IR）不敏感。我们还表明，Snm1B缺陷型细胞在对MMC的反应中表现出有缺陷的S期检查点，但对IR没有，这一发现可能解释了对交联药物的特异性敏感性。有趣的是，尽管先前的研究在很大程度上暗示ATR是响应ICL而被激活的主要激酶，但我们表明在Snm1B缺陷型细胞中存在缺陷的是ATM介导的检查点的激活。在ICL损伤后，ATM检查点激活对Snm1B的需求与其在促进双链断裂形成从而导致复制叉崩溃中的作用相关。与这一结果一致，发现Snm1B直接与Mus81-Eme1相互作用，Mus81-Eme1是一种先前与叉崩溃有关的核酸酶。此外，我们还表明Snm1B与Mre11-Rad50-Nbs1（MRN）复合物以及FancD2相互作用，进一步证实了其作为检查点/DNA修复蛋白的作用。
