DNA Repair and Replication Re-start in vivo

描述(由申请人提供)：DNA损伤及其修复是基因组不稳定的主要因素，而基因组不稳定阻碍了肿瘤的发展。这项建议阐述了一系列在避免癌症中重要的DNA损伤修复蛋白的功能。本项目研究大肠杆菌RecQ DNA解旋酶：一个高度保守的蛋白质家族的创始成员，该家族包括五个人类同源物，三个与癌症易感综合征有关--Bloom、Werner和Rothmund-Thomson。最近研究表明，E.ColiRecQ对细胞的同源重组修复(HR)有全面的影响，而不同于几个研究得很好的RecQ同源物，包括酵母Sgs1、人WRN和可能的BLM。虽然一些RecQ同系物促进HR(分子间重组中间体或IRIS)中分子间中间体在细胞中的净减少，这可以阻止染色体分离，导致死亡，但RecQ可以促进IRIS在体内的净积累。这为RecQ样蛋白的整体体内效应构成了一个新的第二范式，并可能为一些人类同源物的细胞作用提供一个更好的模型。该项目将阐明E.ColiRecQ在体内的功能，并将解决哪些人类同源物的功能相似。这项工作利用了大肠杆菌模型系统无与伦比的可操作性、复杂的遗传和其他工具，以及它作为第一个研究新的RecQ范式的模型系统的独特地位，也是目前唯一也具有适当实验试剂的系统。该项目中的大肠杆菌工作为理解RecQ同源物及其在受损复制叉处的DNA修复中的作用，以及驱动基因组不稳定的自发DNA损伤和修复提供了独特的切入点。人类细胞工作将把这些发现转化为与癌症相关的蛋白质/基因和疾病。两者都将阐明并解释基因组稳定性的保守的基本机制，并指出在探索高度相似的人类途径方面需要追求的新方向。公共卫生相关性：该项目的目标是确定RecQ蛋白家族成员在活细胞中的功能，该家族蛋白构成了一类重要的癌症避免蛋白。这些蛋白质修复DNA，避免基因组不稳定，因此有助于预防癌症。通过研究细菌中的相关蛋白，已经取得了重大进展。在这里，我们将利用这些进展，并确定与大肠杆菌模型中最相似的人类RecQ亲属。这项工作将为理解和诊断、治疗和预防由这些蛋白质功能障碍引起的人类癌症提供基础。

DESCRIPTION (provided by applicant): DNA damage and its repair are the primary contributors to genome instability that underly tumor development. This proposal addresses the function of a family of DNA- damage-repair proteins important in cancer avoidance. This project investigates E. coli RecQ DNA helicase: the founding member of a highly-conserved protein family that includes five human homologues, three associated with cancer-predisposition syndromes--Bloom, Werner, and Rothmund-Thomson. E. coli RecQ was shown recently to exert an overall effect on homologous-recombinational repair (HR) in cells different from several well-studied RecQ homologues including yeast Sgs1 and human WRN and possibly BLM. Whereas some RecQ homologues promote the net reduction in cells of intermolecular intermediates in HR (intermolecular recombination intermediates or IRIs), which can block chromosome segregation causing death if left unresolved, RecQ can promote net accumulation of IRIs in vivo. This constitutes a new, second paradigm for the overall in vivo effect of RecQ-like proteins, and may provide a better model of the cellular role of some of the human homologues. This project will elucidate the function of E. coli RecQ in vivo, and will address which if any human homologues function similarly. This work exploits the unparalleled tractability, sophisticated genetic and other tools of the E. coli model system, and its unique status as the first model system in which to investigate the new RecQ paradigm, and currently the only system that also has appropriate experimental reagents. The E. coli work in this project provides unique entry points into understanding RecQ homologues and their roles in DNA repair at damaged replication forks, and into spontaneous DNA damage and repair that drive genome instability. The human cell work will translate these discoveries to the relevant cancer- related proteins/genes and diseases. Both will illuminate then translate well conserved basic mechanisms of genome (in)stability, and indicate new directions to be pursued in exploring the highly similar human pathways. PUBLIC HEALTH RELEVANCE: The goal of this project is to determine the functions in living cells of the members of the RecQ family of proteins, which constitute an important class of cancer-avoidance proteins. These proteins repair DNA and avoiding genome instability, and so help to prevent cancers. Major advances have occurred by studying the related protein in the bacterium E. coli. Here we will capitalize on those advances and also identify the human RecQ relatives most like that in the E. coli model. This work will provide a foundation for understanding, and so for diagnosis, treatment, and prevention, of human cancers caused by malfunction of these proteins.