The mismatch repair system (mutS, mutL and uvrD genes) in Pseudomonas aeruginosa: molecular characterization of naturally occurring mutants
Related Experiment Video
Updated: Oct 1, 2026
Replication of the Ordered, Nonredundant Library of Pseudomonas aeruginosa strain PA14 Transposon Insertion Mutants
Published on: May 4, 2018
Antonio Oliver1, Fernando Baquero, Jesús Blázquez
1Unidad de Microbiología Molecular, Servicio de Microbiología, Hospital Ramón y Cajal, Carretera de Colmenar Km 9.1, 28034-Madrid, Spain.
Abstract:
We have recently described the presence of a high proportion of Pseudomonas aeruginosa isolates (20%) with an increased mutation frequency (mutators) in the lungs of cystic fibrosis (CF) patients. In four out of 11 independent P. aeruginosa strains, the high mutation frequency was found to be complemented with the wild-type mutS gene from P. aeruginosa PAO1. Here, we report the cloning and sequencing of two additional P. aeruginosa mismatch repair genes and the characterization, by complementation of deficient strains, of these two putative P. aeruginosa mismatch repair genes (mutL and uvrD). We also describe the alterations in the mutS, mutL and uvrD genes responsible for the mutator phenotype of hypermutable P. aeruginosa strains isolated from CF patients. Seven out of the 11 mutator strains were found to be defective in the MMR system (four mutS, two mutL and one uvrD). In four cases (three mutS and one mutL), the genes contained frameshift mutations. The fourth mutS strain showed a 3.3 kb insertion after the 10th nucleotide of the mutS gene, and a 54 nucleotide deletion between two eight nucleotide direct repeats. This deletion, involving domain II of MutS, was found to be the main one responsible for mutS inactivation. The second mutL strain presented a K310M mutation, equivalent to K307 in Escherichia coli MutL, a residue known to be essential for its ATPase activity. Finally, the uvrD strain had three amino acid substitutions within the conserved ATP binding site of the deduced UvrD polypeptide, showing defective mismatch repair activity. Interestingly, cells carrying this mutant allele exhibited a fully active UvrABC-mediated excision repair. The results shown here indicate that the putative P. aeruginosa mutS, mutL and uvrD genes are mutator genes and that their alteration results in a mutator phenotype.
Insights
Mutations in mismatch repair genes (mutS, mutL, uvrD) cause a mutator phenotype in Pseudomonas aeruginosa from cystic fibrosis patients. Defective mismatch repair (MMR) contributes to increased mutation frequency in these bacteria.
Area of Science:
- Microbiology
- Genetics
- Molecular Biology
Background:
- Pseudomonas aeruginosa is a significant opportunistic pathogen, particularly in cystic fibrosis (CF) patients.
- A high proportion (20%) of P. aeruginosa isolates from CF patients exhibit a mutator phenotype, characterized by increased mutation frequency.
- Previous work identified mutS gene complementation in some hypermutable strains.
Purpose of the Study:
- To clone, sequence, and characterize additional mismatch repair (MMR) genes (mutL and uvrD) in P. aeruginosa.
- To identify specific alterations in mutS, mutL, and uvrD responsible for the mutator phenotype in CF isolates.
- To confirm the role of these genes in the MMR system of P. aeruginosa.
Main Methods:
- Cloning and sequencing of P. aeruginosa mutL and uvrD genes.
- Complementation assays using deficient P. aeruginosa strains.
- Analysis of gene alterations (frameshift mutations, insertions, deletions, amino acid substitutions) in mutS, mutL, and uvrD.
- Assessment of mismatch repair and excision repair activities.
Main Results:
- Seven of 11 mutator strains were defective in the MMR system, including four with mutS, two with mutL, and one with uvrD defects.
- Specific mutations identified include frameshift mutations, a large insertion and deletion in mutS, a K310M substitution in mutL affecting ATPase activity, and three substitutions in the UvrD ATP binding site.
- The identified mutations in mutS, mutL, and uvrD were responsible for the mutator phenotype.
- The uvrD mutant allele did not impair UvrABC-mediated excision repair.
Conclusions:
- The P. aeruginosa mutS, mutL, and uvrD genes are crucial for maintaining genomic stability.
- Alterations in these MMR genes lead to a mutator phenotype in P. aeruginosa strains from CF patients.
- Defective MMR is a significant factor contributing to the genetic adaptability and potential virulence of P. aeruginosa in the CF lung environment.
More Related Videos
08:57Identification of Novel Genes Associated with Alginate Production in Pseudomonas aeruginosa Using Mini-himar1 Mariner Transposon-mediated Mutagenesis
Published on: March 10, 2014
08:34Generation of In-Frame Gene Deletion Mutants in Pseudomonas aeruginosa and Testing for Virulence Attenuation in a Simple Mouse Model of Infection
Published on: January 8, 2020
Related Concept Videos
01:48Mismatch Repair
01:20Mismatch Repair
The Mutator Protein Family Plays a Key Role in DNA Mismatch Repair
The human genome has more than 3 billion base pairs of DNA per cell. Prior to cell division, that vast amount of genetic...
01:48Mismatch Repair
01:18Other Unique Bacteria
01:38Nucleotide Excision Repair
Cells are regularly exposed to mutagens—factors in the environment that can damage DNA and generate mutations. UV radiation is one of the most common mutagens and is estimated to introduce a significant number of changes in DNA. These include bends or kinks in the structure, which can block DNA replication or transcription. If these errors are not fixed, the damage can cause mutations, which in turn can result in cancer or disease depending on which sequences are...
01:46Nucleotide Excision Repair