Role of HELLS chromatin remodeler in genome maintenance

项目摘要/母基金摘要（1 R 01 GM 143428 -01） 稳健和严格调控的DNA修复对于维持基因组稳定性和预防疾病至关重要 发展真核DNA被包装成染色质，染色质具有深刻的，但尚未充分理解的调控作用。 影响DNA修复、复制和基因组维持。在理解上存在根本性的差距 DNA修复途径是如何在结构多样的染色质中进行调节和协调的， 异质基因组景观。HELLS（Helicase cytophoid Specific）是一种知之甚少的染色质， 相关蛋白，在基因组维护中具有新兴的新作用。鼠标地狱是必不可少的 配子发生和免疫系统的正常发育。人类HELLS的突变导致严重的 免疫缺陷综合征ICF（免疫缺陷着丝粒不稳定性面部异常）。尽管 在理解哺乳动物HELLS在DNA甲基化中的分子功能方面取得了重大进展， 染色质重塑，其在DNA修复和基因组维护中的作用知之甚少，仍然难以捉摸。 未解决的问题仍然是，HELLS是否调节多个DNA修复途径，以及它是否 在修复和维持不同的基因组基因座或结构域中的专门作用。我们已经建立和 经过验证的真菌模型，粗糙脉孢菌，以推进对HELL介导的 基因组稳定性的机制。我们的研究揭示了一个新的，以前未被认识的地狱之间的联系 真菌和人类细胞中蛋白质和细胞对DNA烷基化损伤的反应。我们假设 HELLS保护细胞免受烷基化诱导的毒性，并在细胞的修复和稳定中发挥重要作用。 组成型异染色质结构域。这一假设是建立在一个强有力的初步数据的基础上， 在真菌模型脉孢菌和人类细胞中，证明缺乏HELLS的细胞表现出对 DNA烷基化损伤和组成性异染色质修复缺陷。另外我们 发现在HELLS突变体细胞中真菌WDR 76蛋白的丢失导致合成性的拯救。 烷基化敏感性表型，这意味着WDR 76作为HELLS缺陷的遗传抑制因子。目标1 我们将确定真菌和人类HELLS重塑者在烷基化DNA损伤修复中的作用。在 目标2我们将定义精确的基因组和染色质的背景下，依赖于地狱的基因组维护。 在目标3中，我们将确定HELLS和WDR 76之间的功能关系，WDR 76是一种涉及WD 40的蛋白质， 对DNA烷基化的反应。成功完成拟议的研究将定义地狱介导的 基因组维护的机制，并确定其他监管机构和途径与地狱合作 保护细胞免受遗传毒性应激的有害后果。这些研究将提供重要的 深入了解致病染色体重排和断裂的起源，在许多人中发现， 疾病，包括ICF综合征。

Project summary/abstract from parent grant (1R01GM143428-01) Robust and tightly regulated DNA repair is critical to maintain genome stability and prevent disease development. Eukaryotic DNA is packaged into chromatin that has a profound, yet not well understood regulatory influence on DNA repair, replication, and genome maintenance. There is a fundamental gap in understanding how DNA repair pathways are regulated and coordinated within structurally diverse chromatin, and across the heterogeneous genomic landscape. The HELLS (Helicase Lymphoid Specific) is a poorly understood chromatin- associated protein, with an emerging new role in genome maintenance. Mouse HELLS is essential for gametogenesis and proper development of the immune system. Mutations in human HELLS cause severe immunodeficiency syndrome ICF (Immunodeficiency Centromeric Instability Facial anomalies). Despite substantial progress in understanding the molecular functions of the mammalian HELLS in DNA methylation and chromatin remodeling, its role in DNA repair and genome maintenance is poorly understood and remains elusive. The unresolved questions remain whether HELLS regulates multiple DNA repair pathways, and whether it has specialized roles in the repair and maintenance of a distinct genomic loci or domains. We have established and validated fungal model, Neurospora crassa to advance the fundamental understanding of HELLS-mediated mechanisms of genome stability. Our studies reveal a new, previously unrecognized link between HELLS proteins and cellular responses to DNA alkylation damage in fungal and human cells. We hypothesize that HELLS protects cells form alkylation-induced toxicity and plays important roles in the repair and stability of the constitutive heterochromatin domains. This hypothesis is founded based on a strong preliminary data in the fungal model Neurospora and in human cells demonstrating that cells deficient in HELLS exhibit sensitivity to DNA alkylation damage and are deficient in the repair of the constitutive heterochromatin. In addition, we discovered that loss of fungal WDR76 protein in HELLS mutant cells leads to the synthetic rescue of the alkylation sensitivity phenotype, implying that WDR76 acts as genetic suppressor of HELLS deficiency. In Aim 1 we will determine the role of fungal and human HELLS remodelers in the repair of alkylation DNA damage. In Aim 2 we will define precise genomic and chromatin contexts that depend on HELLS for genome maintenance. In Aim 3 we will determine the functional relationship between HELLS and WDR76, a WD40 protein implicated in response to DNA alkylation. Successful completion of the proposed research will define HELLS-mediated mechanisms of genome maintenance, and identify additional regulators and pathways cooperating with HELLS in protecting the cells from detrimental consequences of genotoxic stress. These studies will provide important insights into the origin of the disease-causing chromosomal rearrangements and breaks found in many human diseases, including the ICF syndrome.