BACKGROUND Acinetobacter baylyi ADP1 is an ideal bacterial strain for high-throughput genetic analysis as the bacterium is naturally transformable. Thus, ADP1 can be used to investigate DNA mismatch repair, a mechanism for repairing mismatched bases. We used the mutS deletion mutant (XH439) and mutL deletion mutant (XH440), and constructed a mutS mutL double deletion mutant (XH441) to investigate the role of the mismatch repair system in A. baylyi. RESULTS We determined the survival rates after UV irradiation and measured the mutation frequencies, rates and spectra of wild-type ADP1 and mutSL mutant via rifampin resistance assay (RifR assay) and experimental evolution. In addition, transformation efficiencies of genomic DNA in ADP1 and its three mutants were determined. Lastly, the relative growth rates of the wild type strain, three constructed deletion mutants, as well as the rifampin resistant mutants obtained from RifR assays, were measured. All three mutants had higher survival rates after UV irradiation than wild type, especially the double deletion mutant. Three mutants showed higher mutation frequencies than ADP1 and favored transition mutations in RifR assay. All three mutants showed increased mutation rates in the experimental evolution. However, only XH439 and XH441 had higher mutation rates than the wild type strain in RifR assay. XH441 showed higher transformation efficiency than XH438 when donor DNA harbored transition mutations. All three mutants showed higher growth rates than wild-type, and these four strains displayed higher growth rates than almost all their rpoB mutants. The growth rate results showed different amino acid mutations in rpoB resulted in different extents of reduction in the fitness of rifampin resistant mutants. However, the fitness cost brought by the same mutation did not vary with strain background. CONCLUSIONS We demonstrated that inactivation of both mutS and mutL increased the mutation rates and frequencies in A. baylyi, which would contribute to the evolution and acquirement of rifampicin resistance. The mutS deletion is also implicated in increased mutation rates and frequencies, suggesting that MutL may be activated even in the absence of mutS. The correlation between fitness cost and rifampin resistance mutations in A. baylyi is firstly established.

中文翻译：

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贝氏不动杆菌ADP1中的错配修复系统（mutS和mutL）。

背景技术Baylyi不动杆菌ADP1是用于高通量遗传分析的理想细菌菌株，因为该细菌是天然可转化的。因此，ADP1可用于研究DNA错配修复，这是一种修复错配碱基的机制。我们使用了mutS缺失突变体（XH439）和mutL缺失突变体（XH440），并构建了一个mutS mutL双缺失突变体（XH441），以研究错配修复系统在Baylyi中的作用。结果我们确定了紫外线照射后的存活率，并通过利福平抗性测定（RifR测定）和实验演变来测量野生型ADP1和mutSL突变体的突变频率，突变率和光谱。另外，测定了ADP1及其三个突变体中基因组DNA的转化效率。最后，野生型菌株的相对生长速率 测量了三个构建的缺失突变体，以及从RifR分析获得的耐利福平的突变体。这三个突变体在紫外线照射后的存活率均高于野生型，尤其是双缺失突变体。三个突变体显示出比ADP1高的突变频率，并且在RifR分析中倾向于过渡突变。所有三个突变体在实验进化中均显示出增加的突变率。但是，在RifR分析中，只有XH439和XH441具有比野生型菌株更高的突变率。当供体DNA具有过渡突变时，XH441显示出比XH438更高的转化效率。所有三个突变体均显示出比野生型更高的生长速率，并且这四个菌株均比几乎所有rpoB突变体均显示出更高的生长速率。增长率结果表明，rpoB中不同的氨基酸突变导致耐利福平突变体的适应性降低程度不同。但是，同一突变带来的适应性成本并未随菌株背景而变化。结论我们证明mutS和mutL的失活增加了A. baylyi的突变率和频率，这将有助于利福平耐药性的演变和获得。mutS缺失也与突变率和频率增加有关，这表明即使在不存在mutS的情况下，MutL也可能被激活。首先建立了A.baylyi适合度成本与利福平抗性突变之间的相关性。相同突变带来的适应性成本不随菌株背景而变化。结论我们证明mutS和mutL的失活增加了A. baylyi的突变率和频率，这将有助于利福平耐药性的演变和获得。mutS缺失也与突变率和频率增加有关，这表明即使在不存在mutS的情况下，MutL也可能被激活。首先建立了A.baylyi适合度成本与利福平抗性突变之间的相关性。相同突变带来的适应性成本不随菌株背景而变化。结论我们证明mutS和mutL的失活增加了A. baylyi的突变率和频率，这将有助于利福平耐药性的演变和获得。mutS缺失也与突变率和频率增加有关，这表明即使在不存在mutS的情况下，MutL也可能被激活。首先建立了A.baylyi适合度成本与利福平抗性突变之间的相关性。mutS缺失也与突变率和频率增加有关，这表明即使在不存在mutS的情况下，MutL也可能被激活。首先建立了A.baylyi适合度成本与利福平抗性突变之间的相关性。mutS缺失也与突变率和频率增加有关，这表明即使在不存在mutS的情况下，MutL也可能被激活。首先建立了A.baylyi适合度成本与利福平抗性突变之间的相关性。