Role of HELLS chromatin remodeler in genome maintenance

项目摘要/母基金摘要(1R01GM143428-01) 强大和严格调控的DNA修复是维持基因组稳定和预防疾病的关键 发展。真核DNA被包装成染色质，它具有一种深刻但尚未被很好理解的调控 对DNA修复、复制和基因组维护的影响。在理解上存在着根本性的差距 DNA修复通路是如何在结构不同的染色质内以及在 异质基因组景观。Hells(解旋酶淋巴特异的)是一种鲜为人知的染色质- 相关蛋白，在基因组维护中发挥着新的作用。老鼠地狱是必不可少的 配子发生和免疫系统的正常发育。人类地狱中的突变导致严重的 免疫缺陷综合征ICF(免疫缺陷着丝粒不稳定面部畸形)。尽管 了解哺乳动物Hells在DNA甲基化和DNA甲基化中的分子功能取得实质性进展 染色质重塑，它在DNA修复和基因组维持中的作用还知之甚少，仍然难以捉摸。 悬而未决的问题仍然是，地狱是否调控了多条DNA修复途径，以及是否调控了 在修复和维护不同的基因组位点或结构域中的特殊作用。我们已经建立了 验证真菌模型粗糙脉孢菌促进对Hells介导的基础理解 基因组稳定性的机制。我们的研究揭示了地狱之间一种新的、以前未被认识到的联系 真菌和人类细胞对DNA烷基化损伤的蛋白质和细胞反应。我们假设 Hells保护细胞免受烷基化毒性，在细胞修复和稳定中发挥重要作用。 构成异染色质结构域。这一假设是建立在强大的初步数据基础上的 真菌模型神经孢子虫和在人类细胞中表明地狱缺陷的细胞对 DNA烷基化损伤，缺乏对构成异染色质的修复。此外，我们 发现在Hells突变细胞中真菌WDR76蛋白的丢失导致了对 烷基化敏感性表型，暗示WDR76是Hells缺乏症的遗传抑制基因。在目标1中 我们将确定真菌和人类地狱重构体在烷基化DNA损伤修复中的作用。在……里面 目的2我们将定义依赖于地狱来维持基因组的精确的基因组和染色质环境。 在目标3中，我们将确定Hells和WDR76之间的功能关系，WD40蛋白与WD40有关 作为对DNA烷基化的反应。成功完成拟议的研究将定义地狱调解 基因组维持的机制，并确定与地狱合作的其他调节因子和途径 在保护细胞免受遗传毒性压力的有害后果方面。这些研究将提供重要的 对许多人类中发现的致病染色体重排和断裂的起源的洞察 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.