Probing the molecular function of RecQ4 in human Genome Integrity

探讨 RecQ4 在人类基因组完整性中的分子功能

• 批准号: 396699286
• 负责人: Professorin Dr. Caroline Kisker
• 金额: --
• 依托单位: Rudolf-Virchow-Zentrum - Center for Integrative and Translational Bioimaging
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2018
• 资助国家: 德国
• 起止时间: 2017-12-31 至 2022-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/396699286?language=en
• 关键词: Probing; molecular; function; RecQ4; human

Maintenance of genomic integrity is vital for every organism to preserve cellular functions and prevent cancer and aging. The family of RecQ helicases represents an essential component of the genome maintenance system. Its members operate in virtually all DNA metabolism pathways, including DNA replication, DNA repair and telomere maintenance. Of the five RecQ helicases found in humans, RecQ4 assumes a special position as it lacks several RecQ characteristics domains, which have been shown to be indispensible for functionality in other RecQ helicases. Furthermore, RecQ4 is the only RecQ family member which is also present in mitochondria in addition to the nucleus. Loss of function mutations in RecQ4 lead to the development of the three human pathologies Rothmund Thomson Syndrome type II, RAPADILINO syndrome and Baller Gerold Syndrome, characterized by a combination of developmental defects, skeletal abnormalities, a predisposition to cancer and/or premature aging phenotypes. Intriguingly, RecQ4 upregulation is also associated with carcinogenesis and metastasis. We solved the structure of the RecQ4 helicase and showed that it contains a unique domain within its C terminus that likely represents a module for the interaction with other proteins and/or specific non standard DNA substrates. Our biochemical data indicate that RecQ4 might utilize an atypical helicase mechanism, whose function is facilitated by the very C terminal amino acids. In order to elucidate the molecular basis of this uncommon mechanism we will structurally and functionally characterize RecQ4 with its C terminus in complex with different DNA substrates. To decipher the functional role of our newly discovered domain, we will conduct binding studies with non standard DNA substrates, such as Holiday Junctions, Displacement loops and G quadruplex DNA, all of which resemble naturally occurring DNA metabolism intermediates, and examine if and how RecQ4 could process such DNA structures. The molecular framework of how RecQ4 facilitates genomic maintenance is only beginning to emerge. Thus, we will structurally and functionally characterize the binding between RecQ4 and three important DNA maintenance related binding partners: PARP 1, p53 and XPA. After determining the minimal interaction complexes, which will be utilized for structural studies, we will probe the functional collaboration between the binding partners, both in vitro and in vivo. The analysis of the atypical helicase mechanism of RecQ4 in combination with functional interaction studies addressing important genome maintenance related DNA substrates as well as binding partners will provide important insights into the role of RecQ4 in genome maintenance and the origin of RecQ4 associated human pathologies.

维持基因组完整性对每个生物体保持细胞功能、预防癌症和衰老至关重要。RecQ解旋酶家族是基因组维持系统的重要组成部分。它的成员几乎参与了所有的DNA代谢途径，包括DNA复制、DNA修复和端粒维护。在人类发现的五种RecQ解旋酶中，RecQ4具有特殊的位置，因为它缺乏几个RecQ特征结构域，这些结构域已被证明是其他RecQ解旋酶功能所必需的。此外，RecQ4是RecQ家族成员中唯一存在于线粒体和细胞核中的。RecQ4的功能突变缺失导致了Rothmund Thomson综合征II型、RAPADILINO综合征和Baller Gerold综合征三种人类病理的发展，其特征是发育缺陷、骨骼异常、易患癌症和/或早衰表型的组合。有趣的是，RecQ4上调也与癌变和转移有关。我们解决了RecQ4解旋酶的结构，并表明它在其C端包含一个独特的结构域，可能代表了与其他蛋白质和/或特定非标准DNA底物相互作用的模块。我们的生化数据表明，RecQ4可能利用非典型解旋酶机制，其功能是由C端氨基酸促进的。为了阐明这种罕见机制的分子基础，我们将对RecQ4及其C端与不同DNA底物的复合物进行结构和功能表征。为了破译我们新发现的结构域的功能作用，我们将对非标准DNA底物进行结合研究，如假日连接，位移环和G四重DNA，所有这些都类似于自然发生的DNA代谢中间体，并检查RecQ4是否以及如何处理这些DNA结构。RecQ4如何促进基因组维持的分子框架才刚刚开始出现。因此，我们将从结构和功能上表征RecQ4与三个重要的DNA维持相关的结合伙伴：parp1、p53和XPA之间的结合。在确定了用于结构研究的最小相互作用复合物后，我们将在体外和体内探索结合伙伴之间的功能协作。对RecQ4非典型解旋酶机制的分析，结合针对重要基因组维持相关DNA底物和结合伙伴的功能相互作用研究，将为了解RecQ4在基因组维持中的作用以及RecQ4相关人类病理的起源提供重要的见解。