Translesion synthesis in Escherichia coli: Lessons from the NarI mutation hot spot
Translesion synthesis in Escherichia coli: Lessons from the NarI mutation hot spot
复制标题
DOI：
10.1016/j.dnarep.2007.02.021
复制
发表时间：
2007-07-01
期刊：
影响因子：
3.8
通讯作者：
Fujii, Shingo
中科院分区：
文献类型：
--
作者：
Fuchs, Robert P.;Fujii, Shingo
含有受损碱基的DNA的复制对于通常仅以高速度、高准确性和高持续合成能力的未受损模板复制的DNA聚合酶是一个挑战。当一个复制型DNA聚合酶遇到一个化学改变的碱基，它不能复制，一个过程称为translesion合成（TLS）发生，在此期间，复制型聚合酶被一个所谓的专门或病变旁路聚合酶瞬时取代。除了作为复制和跨损伤DNA聚合酶的中心参与者之外，TLS途径还涉及辅助因子，例如一般复制持续合成因子（即原核生物中的β-夹和真核生物中的PCNA）。在大肠杆菌中，除了β-夹，RecA作为Pol V的辅因子在该生物体中发挥重要作用，Pol V是主要的旁路聚合酶。TLS途径的综合观点必然需要遗传和生化研究。在这篇综述中，我们将试图总结的见解TLS在过去的25年中获得了研究移码突变的热点，NarI网站。该位点最初是在建立由化学致癌物N-2-乙酰氨基芴（AAF）诱导的正向突变谱时偶然发现的。事实上，这种化学致癌物与DNA共价结合，形成与鸟嘌呤残基的加合物。当与NarI位点的G* 结合时，5 '-GGCG*CC-，AAF以比自发频率高约10（7）倍的频率诱导G*pC二核苷酸的损失。体内研究表明，NarI突变热点既不限于NarI序列本身，也不限于致癌物AAF。相反，热点需要含有至少两个GpC重复序列和形成一大类人类致癌物的芳香酰胺和硝基芳香族化合物家族中的任何一种的序列。遗传分析最初揭示，NarI移码途径是SOS依赖性的，但不依赖于umuDC（即Pol V）。最近，DNA Pol II被鉴定为负责这种移码途径的酶。同时，AAF加合物在NarI网站可以绕过在一个错误的方式由Pol V. NarI网站，因此提供了一个独特的可能性，研究两个专门的DNA聚合酶，Pol II和Pol V之间的相互作用，既可以延长复制中间体形成时，复制Pol III解离在附近的损害。这两种途径的完全重建使我们强调了TLS途径的一个关键特征，即在单一结合事件的过程中，一片DNA合成（TLS补丁），其足够长，以便在重新加载复制DNA聚合酶时从校正活动中“隐藏损伤诱导的失真”（或当专门的DNA聚合酶分离时可以接近引物的任何核酸外切酶）。所有DNA聚合酶都与之结合的β-夹在允许专门的DNA聚合酶合成足够长的TLS补丁以抵抗这种“外部校对”活动方面起着关键作用。(C)2007 Elsevier B. V.保留所有权利。
Duplication of DNA containing damaged bases is a challenge to DNA polymerases that normally replicate with high speed, high accuracy and high processivity undamaged templates only. When a replicative DNA polymerase encounters a chemically altered base that it is unable to copy, a process called translesion synthesis (TLS) takes place during which the replicative polymerase is transiently replaced by a so-called specialized or lesion bypass polymerase. in addition to the central players that are the replicative and translesion DNA polymerases, TLS pathways involve accessory factors such as the general replication processivity factor (i.e. the beta-clamp in prokaryotes and PCNA in eukaryotes). In Escherichia coli, besides the beta-clamp, RecA plays a fundamental role as a co-factor of Pol V the major bypass polymerase in this organism. An integrated view of TLS pathways necessarily requires both genetic and biochemical studies. In this review we will attempt to summarize the insights into TLS gained over the last 25 years by studying a frameshift mutation hot spot, the NarI site. This site was initially discovered by serendipity when establishing a forward mutation spectrum induced by a chemical hepatocarcinogen, N-2-acetylaminofluorene (AAF). Indeed, this chemical carcinogen covalently binds to DNA forming adducts with guanine residues. When bound to G* in the NarI site, 5'-GGCG*CC-, AAF induces the loss of the G*pC dinucleotide at a frequency that is approximate to 10(7)-fold higher than the spontaneous frequency. In vivo studies showed that the NarI mutation hot spot is neither restricted to the NarI sequence itself, nor to the carcinogen AAF. Instead, the hot spot requires a sequence containing at least two GpC repeats and any of a family of aromatic amides and nitro aromatic compounds that form a large class of human carcinogens. Genetic analysis initially revealed that the NarI frameshift pathway is SOS dependent but umuDC (i.e. Pol V) independent. More recently, DNA Pol II was identified as the enzyme responsible of this frameshift pathway. Concurrently the AAF adduct in the NarI site can be bypassed in an error-free way by Pol V. The NarI site thus offers a unique possibility to study the interplay between two specialized DNA polymerases, Pol II and Pol V, that can both extend replication intermediates formed when the replicative Pol III dissociates in the vicinity of the damage. Full reconstitution of the two pathways led us to highlight a key feature for TLS pathways, namely that it is critical the specialized DNA polymerase synthesizes, during the course of a single binding event, a patch of DNA synthesis (TLS patch) that is long enough as to "hide the lesion induced distortion" from the proofreading activity upon reloading of the replicative DNA polymerase (or any exonuclease that may get access to the primer when the specialized DNA polymerase detaches). The beta-clamp, to which all DNA polymerases bind, plays a critical role in allowing the specialized DNA polymerases to synthesize TLS patches that are long enough to resist such "external proofreading" activities. (C) 2007 Elsevier B.V. All rights reserved.