The role of And-1 in nucleotide excision repair

项目摘要 核苷酸切除修复(NER)是去除紫外线引起的DNA大损伤的主要途径， 环境诱变剂和化疗药物。与NER相关的基因缺失已被联系起来 到着色性干皮病(XP)和皮肤癌。检测NER对DNA的损伤有两种机制， 一种是全球基因组NER(GG-NER)，另一种是转录偶联NER(TC-NER)。GG-NER出现 基因组中的任何位置，而TC-NER负责加速修复 转录的活性基因链。这两条道路都分为早期阶段和晚期阶段。最初的一步是 XP蛋白识别、解开和切割DNA损伤的一系列动作。后一步是 在这两种机制上是相同的，其特征是缺口填充修复合成，其中DNA复制 蛋白质填充~30个核苷酸的缺口，然后连接。与特征明确的早期步骤相比， 调控晚期缝隙DNA合成激活的分子机制仍很大程度上仍然存在 未知。对这种调节途径的生理影响更是知之甚少。 And-1是一种酸性核质DNA结合蛋白，其酵母同源基因Ctf4最初是 被确认为染色体稳定性的关键基因。有趣的是，Ctf4基因缺失的酵母细胞 对紫外线高度敏感，提示和-1在紫外线诱导的DNA损伤反应中起作用。然而，如何 而-1对NER的调节在很大程度上仍是未知的。在这项研究中，我们现在有了广泛的初步数据 证明And-1通过调节缝隙填充DNA的合成而对NER起关键作用。我们的假设是 和-1调节紫外线损伤部位的DNA聚合酶活性，以激活晚期填补缺口的DNA合成 NER的阶段。为了验证这一假设，我们计划追求三个具体目标。目标1：确定机制 And-1通过其调节NER中DNA聚合酶的活性。目标2：确定独一无二的机制 和-1被招募到紫外线损伤部位。目的3：确定和-1在NER和皮肤肿瘤发生中的作用 和-1缺陷小鼠模型。拟议研究的完成不仅将推动该领域的进步 揭示了一种新的和-1介导的调控NER的途径，也为我们提供了体内证据 阐明AND-1在NER和皮肤肿瘤中的新作用。

Project Summary Nucleotide excision repair (NER) is the major pathway to remove bulky DNA lesions induced by UV irradiation, environmental mutagens, and chemotherapeutic agents. Deficiency of genes involved in NER has been linked to Xeroderma Pigmentosum (XP) and skin cancer. There are two mechanisms to detect DNA damage by NER, one is global genome NER(GG-NER) and another is transcription-coupled NER(TC-NER). GG-NER occurs anywhere in the genome, whereas TC-NER is responsible for the accelerated repair of lesions in the transcribed strand of active genes. The both pathways are divided into early and late steps. The early step is the sequential actions, in which XP proteins recognize, unwind, and incise the DNA lesion. The latter step is identical in both mechanisms and is characterized by gap-filling repair synthesis, in which DNA replication proteins fill in the ~30 nucleotide gap, followed by ligation. Compared to the well-characterized early step, the molecular mechanism regulating the activation of gap-filling DNA synthesis at late step remains largely unknown. Even less is known about the physiological impact of such a regulatory pathway. And-1 is an acidic nucleoplasmic DNA-binding protein and its yeast ortholog, Ctf4, was originally identified as a critical gene for chromosome stability. Interestingly, yeast cells with depletion of Ctf4 gene are hypersensitive to UV lights, suggesting a role of And-1 in UV-induced DNA damage response. However, how And-1 regulates NER remains largely unknown. In this study, we now have extensive preliminary data demonstrating that And-1 is critical for NER by regulating gap-filling DNA synthesis. Our hypothesis is that And-1 regulates DNA polymerase activity at UV-lesion sites to activate gap-filling DNA synthesis at the late stage of NER. To test this hypothesis, we plan to pursue three specific aims. Aim 1: Determine the mechanism by which And-1 regulates DNA polymerase activity in NER. Aim 2: Determine the unique mechanism by which And-1 is recruited to UV-lesion sites. Aim 3: Determine the role of And-1 in NER and skin tumorigenesis using And-1 deficient mouse models. The completion of proposed studies will not only advance the field by uncovering a novel And-1-mediated pathway to regulate NER, but also provide us with the in vivo evidence to elucidate a novel role of And-1 in NER and skin tumor.