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Biochemical Investigation on the Mechanism of Error Free Postreplication Repair (PRR) using baker yeast Saccharomyces cerevisiae as a model organism

以面包酵母酿酒酵母为模式生物研究无错复制后修复（PRR）机制的生化研究

基本信息

批准号：
201960739
负责人：
Dr. Christopher Ede
金额：
--
依托单位：
Department of Microbiology and Molecular Genetics
依托单位国家：
德国
项目类别：
Research Fellowships
财政年份：
2011
资助国家：
德国
起止时间：
2010-12-31 至 2012-12-31
项目状态：
已结题

来源：
https://gepris.dfg.de/gepris/projekt/201960739?language=en
关键词：
Biochemical Investigation Mechanism Error Free

项目摘要

DNA lesions can result in an arrest of the replicative DNA-polymerase during replication, which would lead to cell death in case it remains unresolved. Regarding the recent doctrine, three independent processes exist to resolve replication arrests: 1. Translesion synthesis by a specialized DNA-Polymerase (TLS), 2. Error free lesion bypass by homologous recombination with the sister chromatid and 3. Error free Postreplication Repair (PRR) whose mechanisms is still unkown. Usage of these mechanisms is regulated through modification of the processivity factor PCNA: mono-ubiquitylation of lysine 164 results in translesion synthesis, poly-ubiquitylation of lysine 164 results in error free PRR. SUMOylation of lysine 164 instead represses the usage of HR. Recently, data was shown suggesting that HR yet is participating in error free PRR. Regarding this, a new model for error free PRR may be proposed including HR. This model is based upon a dual mechanism: one the one hand, lesion bypass by HR acts as an alternative, in case translesion synthesis proves impossible or inefficient. On the other hand, inefficient TLS by Pol eta may be enhanced by recruiting a second Polymerase (Pol zeta) which supports Pol eta in TLS. Based on this model, two predictions may be made: 1. Efficiency of HR may be enhanced by poly-ubiquitylated PCNA and 2. Efficiency of Pol zeta may be enhanced by poly-ubiquitylated PCNA or Pol zeta is recruited specifically to poly-ubiquitylated PCNA. Aim of the project proposed here is to test these predictions to verify the new model by using biochemical approaches.

DNA损伤可能会导致复制过程中复制的DNA聚合酶停止，这将导致细胞死亡，如果它仍然没有解决。根据最近的学说，存在三个独立的过程来解决复制抑制：1.通过特殊的DNA聚合酶(TLS)合成跨病变；2.通过与姐妹染色单体的同源重组来绕过无错病变；3.无错复制后修复(PRR)，其机制尚不清楚。这些机制的使用是通过修饰过程性因子增殖细胞核抗原来调节的：赖氨酸164的单一泛素化导致跨损伤合成，赖氨酸164的多泛素化导致无错误的PRR。相反，赖氨酸164的总甲基化抑制了HR的使用。最近，有数据显示，HR仍在参与无差错PRR。对此，可以提出一种新的考虑HR的无差错PRR模型。这个模型是基于双重机制的：一方面，在跨病变合成被证明是不可能或无效的情况下，HR的病变旁路可作为替代方案。另一方面，Pol ETA的低效TLS可以通过在TLS中招募第二个支持Pol ETA的聚合酶(Pol Zeta)来增强。基于这个模型，可以做出两个预测：1.多泛素化的增殖细胞核抗原可以提高HR的效率；2.多泛素化的增殖细胞核抗原可以提高Pol Zeta的效率，或者Pol Zeta被特异性地招募到多泛素化的增殖细胞核抗原中。这里提出的项目的目的是检验这些预测，以通过使用生化方法来验证新的模型。