Functional links between Fanconi anemia proteins and interstrand crosslinks

Fanconi贫血(FA)是一种常染色体隐性遗传或X连锁遗传的遗传性疾病。FA患者表现为进行性骨髓功能障碍、全血细胞减少和癌症易感性显著升高。已鉴定出13个不同Fanconi基因(FANCA-I)的种系突变，每个基因对应一个不同的互补组。Fanconi患者细胞的特点是对能够形成DNA链间交联物的双功能烷基化试剂的敏感性显著增加，这表明Fanconi途径在DNA损伤反应中发挥了重要作用。与这一概念一致的是，FANC蛋白的功能似乎围绕着依赖DNA损伤的FANCD2的单一泛素化而组织。FANCI被发现是ATM/ATR检查点信号起始激酶的直接底物。然而，FANC基因产品如何帮助保护细胞免受DNA交联的影响仍然是一个关键问题。项目3的目标是研究招募到DNA链间交联点的蛋白质，并阐明它们在交联链修复和损伤反应信号中的作用。我们的假设是Fanconi贫血途径的组成部分参与了对交联性损伤反应的不同方面。这一假说是通过三个具体目标来解决的。在目标1和2中，我们将使用一种新的方法来研究Fanconi贫血途径的组成部分，以了解它以依赖于DNA复制和不依赖于DNA的方式被招募到交联点的能力。被认为参与ICL加工的候选蛋白质也将检查它们在病变部位的存在。最后，我们将尝试一种无偏见的纯化方法，旨在确定参与交联剂加工的新因素。这些研究有望为了解Fanconi贫血途径以及ICL修复机制提供重要的见解。

Fanconi anemia (FA) is a genetic disorder inherited via autosomal recessive or X-linked patterns. FA patients manifest progressive bone marrow dysfunction, pancytopenia, and drastically elevated cancer predisposition. Germline mutations in 13 different Fanconi genes (FANCA - I) have been identified, each corresponding to a distinct complementation group. The hallmark of Fanconi patient cells is profoundly increased sensitivity to bifunctional alkylating agents capable of forming DNA interstrand crosslinks, suggesting an important role of the Fanconi pathway in DNA damage response. Consistent with this notion, functions of the FANC proteins seems to organize around the DNA damage-dependent monoubiquitination of FANCD2. FANCI was found to be a direct substrate of the ATM/ATR checkpoint signal initiation kinases. However, how FANC gene products help protecting cells from with crosslinked DNA remains a key question. The goal of Project 3 is to study proteins recruited to the site of DNA interstrand crosslinks and to elucidate their role in crosslink repair and damage response signaling. We hypothesis is that components of the Fanconi anemia pathway is involved in different aspects of responses to crosslinking damage. This is hypothesis is addressed by the three Specific Aims. In Aim 1 and 2, we will use a novel approach to investigate components of the Fanconi anemia pathway for its ability to be recruited to the site of crosslink in a DNA replication-dependent and -independent manner. Candidate proteins that have been suggested to have involvement in ICL processing will also be examining for their presence at the site of the lesion. Finally, we will attempt an unbiased purification approach aimed at identify novel factor involved in crosslink processing. These studies are expected to provide significant insights toward the understanding of the Fanconi anemia pathway as well as mechanisms of ICL repair.