Function of RecQ helicases in genome stability

RecQ 解旋酶在基因组稳定性中的功能

• 批准号: 8148298
• 负责人: Vilhelm Bohr
• 金额: $ 106.16万
• 依托单位: NATIONAL INSTITUTE ON AGING
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8148298
• 关键词: Base Excision Repairs; Genome Stability; RecQ4 helicase; helicase; protein complex

In humans there are five RecQ helicases whereas in simpler organisms there is only one. It is of great interest to explore how these RecQ helicases function individually and together and whether there is redundancy of helicase function or whether each has unique individual roles. There may be circumstances where they can function in a synergistic manner and this would be informative about whether these different RecQ helicases interact with each other and also about the biological importance of certain processes. We and other groups find that the human RecQ helicases participate in DNA repair, and more specifically in certain subpathways of DNA repair. Three of the RecQ helicases are deficient in disorders of segmental premature aging: Werner syndrome, Bloom syndrome and Rothmund Thomson syndrome. Much less is known about the Rothmund-Thomson syndrome protein, RECQL4, and RECQL5 than about the other human RecQ helicases. We find that the RECQL4 protein participates in DNA repair of double and single strand breaks and that cells from individuals with this condition are defective in DNA repair. Using real time imaging, we have shown that RECQL4 is recruited to sites of double strand breaks and that its retention kinetics are different than WRN or BLM. Additionally, we mapped the domain of RECQL4 necessary for DNA damage recruitment. Biochemically, we are characterizing the RECQL4 protein and while its helicase function is in many ways similar to WRN and BLM helicases, there are also significant differences. We have shown that RECQL4 has a very limited substrate range and that its helicase activity can be seen on short fork DNA substrates but not on long substrates. We are also exploring potential protein interactions and protein complexes that RECQL4 participates in. We find that RECQL4 can interact with and stimulate the core BER proteins APE1, pol  and FEN1. We are continuing to define and explore the catalytic activities of RECQL4 and potential protein:protein interaction partners. RECQL5 is another member of the RecQ family for which little information is known. We have previously reported that WRN, BLM and RECQL4 can stimulate core base excision repair proteins. Thus we evaluated whether RECQL5 could also stimulate and interact with BER proteins. Indeed, we found that RECQL5 could interact with and stimulate FEN1 incision reactions. We also evaluated RECQL5s ability to interact with and stimulate APE1 or pol however RECQL5 did not stimulate these proteins. Therefore within BER, we are beginning to define which activities are shared among the RecQ helicases and which functions are unique to the individual RecQ helicases.

在人类中有五个RecQ解旋酶，而在更简单的生物体中只有一个。探索这些RecQ解旋酶如何单独和一起发挥作用以及解旋酶功能是否存在冗余或每个解旋酶是否具有独特的个体作用是非常有趣的。可能存在它们可以以协同方式起作用的情况，并且这将提供关于这些不同RecQ解旋酶是否彼此相互作用以及关于某些过程的生物学重要性的信息。我们和其他研究小组发现，人类RecQ解旋酶参与DNA修复，更具体地说，参与DNA修复的某些子途径。RecQ解旋酶中的三种在节段性过早衰老的病症中缺乏：Werner综合征、Bloom综合征和Rothmund Thomson综合征。关于Rothmund-Thomson综合征蛋白RECQL 4和RECQL 5的了解比其他人类RecQ解旋酶少得多。我们发现RECQL 4蛋白参与双链和单链断裂的DNA修复，并且患有这种疾病的个体的细胞在DNA修复中存在缺陷。使用真实的时间成像，我们已经表明，RECQL 4被招募到双链断裂的网站，其保留动力学不同于WRN或BLM。此外，我们绘制了DNA损伤募集所必需的RECQL 4结构域。在生物化学上，我们正在表征RECQL 4蛋白，虽然其解旋酶功能在许多方面与WRN和BLM解旋酶相似，但也存在显着差异。我们已经表明，RECQL 4具有非常有限的底物范围，并且其解旋酶活性可以在短叉DNA底物上看到，但在长底物上看不到。我们还在探索RECQL 4参与的潜在蛋白质相互作用和蛋白质复合物。我们发现RECQL 4可以与BER核心蛋白APE 1、pol和FEN 1相互作用并刺激它们。我们正在继续定义和探索RECQL 4的催化活性和潜在的蛋白质：蛋白质相互作用伙伴。 RECQL 5是RecQ家族的另一个成员，其信息很少。我们以前报道过WRN、BLM和RECQL 4可以刺激核心碱基切除修复蛋白。因此，我们评估了RECQL 5是否也可以刺激BER蛋白并与BER蛋白相互作用。事实上，我们发现RECQL 5可以与FEN 1切口反应相互作用并刺激FEN 1切口反应。我们还评估了RECQL 5与APE 1或pol相互作用并刺激APE 1或pol的能力，但RECQL 5不刺激这些蛋白质。因此，在BER中，我们开始定义RecQ解旋酶之间共享哪些活动，以及哪些功能是单个RecQ解旋酶所独有的。