A novel role for non-ubiquitinated FANCD2 in response to hydroxyurea-induced DNA damage.
A novel role for non-ubiquitinated FANCD2 in response to hydroxyurea-induced DNA damage.
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DOI：
10.1038/onc.2015.68
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发表时间：
2016-01-07
期刊：
影响因子：
8
通讯作者：
Kupfer GM
中科院分区：
文献类型：
--
作者：
Chen X;Bosques L;Sung P;Kupfer GM
范可尼贫血（FA）是一种遗传性疾病的骨髓衰竭，癌症的易感性，并敏感的DNA交联剂。FANCD 2是FA途径的中心蛋白，在DNA损伤后被单泛素化，例如交联剂和复制阻断剂，例如羟基脲（HU）。尽管FA细胞对交联剂如丝裂霉素C（MMC）表现出明确的敏感性，但我们发现它们在很大程度上对羟基脲（HU）具有抗性，除了不表达FANCD 2的细胞。FANCD 2、RAD 51和RAD 18形成复合物，其在HU暴露后增强。令人惊讶的是，虽然FANCD 2是这种增强的相互作用所必需的，但它的单泛素化不是。类似地，非泛素化的FANCD2仍然可以支持PCNA单泛素化。RAD51，而不是BRCA2，也需要PCNA monoubiquitination在响应HU，这表明这一功能是独立的同源重组（HR）。我们进一步表明TLS聚合酶PolH染色质定位在FANCD2缺陷细胞、FANCD2 siRNA敲低细胞和RAD 51 siRNA敲低细胞中降低，并且PolH敲低仅导致HU敏感性。我们的数据表明，FANCD 2和RAD 51在PCNA单泛素化和TLS中起重要作用，FANCD 2单泛素化和HR独立的方式响应HU。MMC处理未观察到这种效应，表明FA途径响应于不同类型的DNA损伤的非典型功能。
Fanconi anemia (FA) is a genetic disease of bone marrow failure, cancer susceptibility, and sensitivity to DNA crosslinking agents. FANCD2, the central protein of the FA pathway, is monoubiquitinated upon DNA damage, such as crosslinkers and replication blockers such as hydroxyurea (HU). Even though FA cells demonstrate unequivocal sensitivity to crosslinkers, such as mitomycin C (MMC), we find that they are largely resistant to hydroxyurea (HU), except for cells absent for expression of FANCD2. FANCD2, RAD51, and RAD18 form a complex, which is enhanced upon HU exposure. Surprisingly, while FANCD2 is required for this enhanced interaction, its monoubiquitination is not. Similarly, non-ubiquitinated FANCD2 can still support PCNA monoubiquitination. RAD51, but not BRCA2, is also required for PCNA monoubiquitination in response to HU, suggesting that this function is independent of homologous recombination (HR). We further show that TLS polymerase PolH chromatin localization is decreased in FANCD2 deficient cells, FANCD2 siRNA knock down cells, and RAD51 siRNA knock down cells, and PolH knockdown results in HU sensitivity only. Our data suggest that FANCD2 and RAD51 play an important role in PCNA monoubiquitination and TLS in a FANCD2 monoubiquitination and HR-independent manner in response to HU. This effect is not observed with MMC treatment, suggesting a non-canonical function for the FA pathway in response to different types of DNA damage.