Regulation of the specialized DNA polymerase eta: Revisiting the biological relevance of its PCNA- and ubiquitin-binding motifs
Regulation of the specialized DNA polymerase eta: Revisiting the biological relevance of its PCNA- and ubiquitin-binding motifs
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DOI：
10.1002/em.21741
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发表时间：
2012-12-01
影响因子：
2.8
通讯作者：
Kannouche, Patricia L.
中科院分区：
文献类型：
--
作者：
Despras, Emmanuelle;Delrieu, Noemie;Kannouche, Patricia L.
在跨损伤合成（TLS）期间，Y家族聚合酶的低保真度聚合酶绕过DNA损伤，所述DNA损伤阻断常规进行性DNA聚合酶的进展，从而允许完成DNA复制。在TLS聚合酶中，DNA聚合酶eta（pol eta）在紫外线（UV）光产物之后进行核苷酸掺入，并且在癌症倾向性着色性干皮病变体（XPV）综合征中缺乏。在UV照射后，DNA滑动钳PCNA在其保守的Lys-164上被单倍酰化。该事件被认为促进了体内TLS过程，因为pol eta通过其泛素结合结构域（UBZ）以及其PCNA相互作用肽（PIP）-盒优先与单泛素化PCNA相互作用。然而，最近的观察对这一模式提出了质疑。因此，在这项研究中，我们重新研究了调节UBZ和PIP结构域的pol eta响应UVC的相对贡献。我们发现，这两个图案的同时失效赋予敏感性UVC，敏化低浓度的咖啡因，DNA合成的长期抑制和持久的S期检查点激活，所有特征的XPV细胞。虽然每个结构域对于pol eta在复制工厂中的有效积累是必不可少的，但UBZ或PIP基序的突变失活仅赋予对UVC的轻微敏感性，这表明尽管信息丰富，但pol eta焦点分析不是评估pol eta在体内TLS中发挥功能的能力的可靠工具。两者合计，这些数据表明，PIP和UBZ基序是不需要的招聘，但保留在网站的停滞复制叉的聚合物。我们认为，这是一种确保有足够量的蛋白质可用于其旁路功能的方法。Environ.摩尔诱变剂，2012. (c)2012 Wiley Periodicals，Inc.
During translesion synthesis (TLS), low-fidelity polymerases of the Y-family polymerases bypass DNA damages that block the progression of conventional processive DNA polymerases, thereby allowing the completion of DNA replication. Among the TLS polymerases, DNA polymerase eta (pol eta) performs nucleotide incorporation past ultraviolet (UV) photoproducts and is deficient in cancer-prone xeroderma pigmentosum variant (XPV) syndrome. Upon UV irradiation, the DNA sliding clamp PCNA is monoubiquitylated on its conserved Lys-164. This event is considered to facilitate the TLS process in vivo since pol eta preferentially interacts with monoubiquitylated PCNA through its ubiquitin-binding domain (UBZ) as well as its PCNA interacting peptide (PIP)-box. However, recent observations questioned this model. Therefore, in this study, we re-examined the relative contribution of the regulatory UBZ and PIP domains of pol eta in response to UVC. We show that simultaneous invalidation of both motifs confers sensitivity to UVC, sensitization by low concentrations of caffeine, prolonged inhibition of DNA synthesis and persistent S phase checkpoint activation, all characteristic features of XPV cells. While each domain is essential for efficient accumulation of pol eta in replication factories, mutational inactivation of UBZ or PIP motif only confers a slight sensitivity to UVC indicating that, although informative, pol eta focus analysis is not a reliable tool to assess the pol eta's ability to function in TLS in vivo. Taken together, these data indicate that PIP and UBZ motifs are not required for recruitment but for retention of pol eta at sites of stalled replication forks. We propose that this is a way to ensure that a sufficient amount of the protein is available for its bypass function. Environ. Mol. Mutagen., 2012. (c) 2012 Wiley Periodicals, Inc.