Survival and induction of SOS in Escherichia coli treated with cisplatin, UV-irradiation, or mitomycin C are dependent on the function of the RecBC and RecFOR pathways of homologous recombination
Survival and induction of SOS in Escherichia coli treated with cisplatin, UV-irradiation, or mitomycin C are dependent on the function of the RecBC and RecFOR pathways of homologous recombination
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DOI：
10.1016/s0921-8777(01)00077-5
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发表时间：
2001-06-05
期刊：
影响因子：
--
通讯作者：
Beck, DJ
中科院分区：
文献类型：
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作者：
Keller, KL;Overbeck-Carrick, TL;Beck, DJ
肿瘤对顺铂和丝裂霉素C（MMC）等药物的耐药性是限制其在癌症化疗中有用性的重要因素。这些药物的抗肿瘤作用是由于在DNA中形成双功能加合物，顺铂主要引起链内交联，MMC引起链间交联。SOS显色试验用于研究暴露于顺铂、紫外线照射（UV）和MMC的大肠杆菌中处理DNA损伤的细胞机制，并随后促进产生诱导SOS反应的分子信号。SOS显色试验中使用的菌株具有lacZ与sfiA（苏拉）基因的融合，从而通过测定β-半乳糖苷酶活性来测量SOS诱导信号的量，所述SOS诱导信号由细胞修复DNA的能力调节。将精通同源重组（HR）的菌株中的SOS诱导与由于recB突变引起的RecBC途径阻断或recO突变引起的RecFOR途径阻断而HR缺陷的等基因菌株中的SOS诱导进行比较。顺铂治疗的效果在一个uvrA突变株阻断在NER的第一步进行了比较，在一个同基因株精通NER。相对于未处理的对照培养物，细胞抗性测量为用增加剂量的顺铂、MMC和UV处理的细胞的集落形成单位（cfu）百分比。两种HR途径对这些处理的抗性的重要性通过突变体中的存活率降低来证明，其中recB突变体比recO突变体更敏感。SOS诱导水平在敏感recB菌株中相对于HR熟练菌株升高，这可能是由于DNA中交联处的停滞和/或扭曲的复制叉。相反，SOS的诱导依赖于RecFOR活性，RecFOR活性被认为作用于新合成的DNA中的子链间隙以介导SOS诱导信号的产生。熟练掌握NER对于顺铂处理的细胞的存活和高水平的SOS诱导都是必要的。(C)出版社：Elsevier Science B. V.
Resistance of tumors to drugs such as cisplatin and mitomycin C (MMC) is an important factor limiting their usefulness in cancer chemotherapy. The antitumor effects of these drugs are due to the formation of bifunctional adducts in DNA, with cisplatin causing predominantly intrastrand-crosslinks and MMC causing interstrand-crosslinks. The SOS chromotest was used to study the cellular mechanisms that process DNA damage in Escherichia coli exposed to cisplatin, ultraviolet irradiation (UV) and MMC and subsequently facilitate the production of a molecular signal for induction of the SOS response. Strains used in the SOS chromotest have a fusion of lacZ with the sfiA (sulA) gene so that the amount of SOS inducing signal, which is modulated by the ability of the cell to repair DNA, is measured by assaying beta -galactosidase activity. SOS induction in a strain proficient in homologous recombination (HR) was compared with that in isogenic strains deficient in HR due to a blocked RecBC pathway caused by a recB mutation or a blocked RecFOR pathway caused by a recO mutation. The effect of cisplatin treatment in a uvrA mutant strain blocked at the first step of NER was compared with that in an isogenic strain proficient in NER. Cellular resistance was measured as percent colony forming units (cfu) for cells treated with increasing doses of cisplatin, MMC and UV relative to that in untreated control cultures. The importance of both HR pathways for resistance to these treatments was demonstrated by decreased survival in mutants with the recB mutant being more sensitive than the recO mutant. SOS induction levels were elevated in the sensitive recB strain relative to the HR proficient strain possibly due to stalled and/or distorted replication forks at crosslinks in DNA. In contrast, induction of SOS was dependent on RecFOR activity that is thought to act at daughter strand gaps in newly synthesized DNA to mediate production of the signal for SOS induction. Proficiency in NER was necessary for both survival and high levels of SOS induction in cisplatin treated cells. (C) 2001 Published by Elsevier Science B.V.