Probing the molecular function of RecQ4 in human Genome Integrity

维持基因组的完整性对于每个生物体保持细胞功能和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。其成员几乎参与所有的DNA代谢途径，包括DNA复制、DNA修复和端粒维持。在人类中发现的五种RecQ解旋酶中，RecQ 4具有特殊的地位，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解旋酶中功能不可或缺的。此外，RecQ 4是除了细胞核之外还存在于线粒体中的唯一RecQ家族成员。RecQ 4中的功能缺失突变导致三种人类病理的发展，即Rothmund Thomson综合征II型、RAPADILINO综合征和Baller格罗尔德综合征，其特征在于发育缺陷、骨骼异常、癌症易感性和/或过早衰老表型的组合。有趣的是，RecQ 4上调也与癌症发生和转移有关。我们解决了RecQ 4解旋酶的结构，并表明它在其C末端包含一个独特的结构域，该结构域可能代表与其他蛋白质和/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，RecQ 4可能利用非典型解旋酶机制，其功能是由非常C末端氨基酸促进。为了阐明这种不常见的机制的分子基础，我们将在结构和功能上表征RecQ 4与其C末端与不同的DNA底物复合。为了破译我们新发现的结构域的功能作用，我们将与非标准DNA底物进行结合研究，如假日接头，置换环和G四链体DNA，所有这些都类似于天然存在的DNA代谢中间体，并检查RecQ 4是否以及如何处理这些DNA结构。RecQ 4如何促进基因组维护的分子框架才刚刚开始出现。因此，我们将在结构和功能上表征RecQ 4与三种重要的DNA维持相关结合伴侣：PARP 1，p53和XPA之间的结合。在确定了最小的相互作用复合物，这将被用于结构研究后，我们将探测结合伙伴之间的功能协作，在体外和体内。RecQ 4的非典型解旋酶机制的分析与功能相互作用研究相结合，解决重要的基因组维护相关的DNA底物以及结合伙伴将提供重要的见解RecQ 4在基因组维护和RecQ 4相关的人类病理学的起源的作用。

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.