SELECTION FOR HIGH MUTATION RATES IN CHEMOSTATS

ABSTRACT Competition experiments in chemostats show that mutator populations of Escherichia coli are more fit than wild type. The increased fitness can be explained by the appearance of new mutants better adapted to the chemostat environment. Fitness values vary between chemostats and are strongly correlated (P < 0.001) with fluctuations in population density.

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REGULATION OF NEWLY EVOLVED ENZYMES. IV. DIRECTED EVOLUTION OF THE EBG REPRESSOR

Genetics ◽

10.1093/genetics/90.4.673 ◽

1978 ◽

Vol 90 (4) ◽

pp. 673-681

Author(s):

Barry G Hall

Keyword(s):

Escherichia Coli ◽

Directed Evolution ◽

Enzyme Synthesis ◽

Regulatory Function ◽

Wild Type ◽

Selection For ◽

Selection Of

ABSTRACT In Escherichia coli, the wild-type repressor of ebg (evolved β-galactosidase) enzyme synthesis, specified by the ebgR + gene, responds very weakly to lactulose (fructose-β-D-galactopyranoside). Selection for a functional repressor that responds strongly to lactulose as an inducer reveals the existence of ebgR+L mutants, which occur spontaneously at a frequency of about 2 x 10-10. ebgR+L mutants are pleiotropic in that they specify ebg repressor with a greatly increased response to lactulose, lactose, galactose-arabinoside and methyl-galactoside as inducers. Selection of ebgR+L mutants is discussed within the framework of directed evolution of a regulatory function.

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Restoration of wild-type motility to flagellin-knockout Escherichia coli

10.1101/700492 ◽

2019 ◽

Author(s):

Nicholas M. Thomson ◽

Mark J. Pallen

Keyword(s):

Escherichia Coli ◽

Copy Number ◽

Plasmid Copy Number ◽

Major Constituent ◽

Strongly Correlated ◽

Wild Type ◽

Plasmid Copy ◽

Inducible Promoters ◽

E Coli ◽

Flagellar Filament

AbstractFlagellin is the major constituent of the flagellar filament and faithful restoration of wild-type motility to flagellin mutants may be beneficial for studies of flagellar biology and biotechnological exploitation of the flagellar system. Therefore, we explored the restoration of motility by flagellin expressed from a variety of combinations of promoter, plasmid copy number and induction strength. Motility was only partially restored using the tightly regulated rhamnose promoter, but wild-type motility was achieved with the T5 promoter, which, although leaky, allowed titration of induction strength. Motility was little affected by plasmid copy number when dependent on inducible promoters. However, plasmid copy number was important when expression was controlled by the native E. coli flagellin promoter. Motility was poorly correlated with flagellin transcription levels, but strongly correlated with the amount of flagellin associated with the flagellar filament, suggesting that excess monomers are either not exported or not assembled into filaments. This study provides a useful reference for further studies of flagellar function and a simple blueprint for similar studies with other proteins.

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Fitness Evolution and the Rise of Mutator Alleles in Experimental Escherichia coli Populations

Genetics ◽

10.1093/genetics/162.2.557 ◽

2002 ◽

Vol 162 (2) ◽

pp. 557-566 ◽

Cited By ~ 5

Author(s):

Aaron C Shaver ◽

Peter G Dombrowski ◽

Joseph Y Sweeney ◽

Tania Treis ◽

Renata M Zappala ◽

...

Keyword(s):

Escherichia Coli ◽

Genetic Drift ◽

Mutation Rates ◽

Significant Gain ◽

Wild Type ◽

Clonal Interference ◽

Beneficial Mutations ◽

Mutator Allele ◽

Experimental Populations ◽

Rate Of Substitution

Abstract We studied the evolution of high mutation rates and the evolution of fitness in three experimental populations of Escherichia coli adapting to a glucose-limited environment. We identified the mutations responsible for the high mutation rates and show that their rate of substitution in all three populations was too rapid to be accounted for simply by genetic drift. In two of the populations, large gains in fitness relative to the ancestor occurred as the mutator alleles rose to fixation, strongly supporting the conclusion that mutator alleles fixed by hitchhiking with beneficial mutations at other loci. In one population, no significant gain in fitness relative to the ancestor occurred in the population as a whole while the mutator allele rose to fixation, but a substantial and significant gain in fitness occurred in the mutator subpopulation as the mutator neared fixation. The spread of the mutator allele from rarity to fixation took >1000 generations in each population. We show that simultaneous adaptive gains in both the mutator and wild-type subpopulations (clonal interference) retarded the mutator fixation in at least one of the populations. We found little evidence that the evolution of high mutation rates accelerated adaptation in these populations.

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ucFabV Requires Functional Reductase Activity to Confer Reduced Triclosan Susceptibility in Escherichia coli

Journal of Molecular Microbiology and Biotechnology ◽

10.1159/000441640 ◽

2015 ◽

Vol 25 (6) ◽

pp. 394-402 ◽

Cited By ~ 1

Author(s):

Taylor L. Fischer ◽

Robert J. White ◽

Katherine F.K. Mares ◽

Devin E. Molnau ◽

Justin J. Donato

Keyword(s):

Escherichia Coli ◽

Reductase Activity ◽

Acid Synthesis ◽

Temperature Sensitive ◽

Wild Type ◽

E Coli ◽

Enoyl Acp Reductase ◽

Selection For

Background/Aims: We previously identified the Triclo1 fosmid in a functional metagenomic selection for clones that increased triclosan tolerance in Escherichia coli. The active enzyme encoded by Triclo1 is ucFabV. Although ucFabV is homologous to FabV from other organisms, ucFabV contains substitutions at key positions that would predict differences in substrate binding. Therefore, a detailed characterization of ucFabV was conducted to link its biochemical activity to its ability to confer reduced triclosan sensitivity. Methods: ucFabV and a catalytic mutant were purified and used to reduce crotonoyl-CoA in vitro. The mutant and wild-type enzymes were introduced into E. coli, and their ability to confer triclosan tolerance as well as suppress a temperature-sensitive mutant of FabI were measured. Results: Purified ucFabV, but not the mutant, reduced crotonoyl-CoA in vitro. The wild-type enzyme confers increased triclosan tolerance when introduced into E. coli, whereas the mutant remained susceptible to triclosan. Additionally, wild-type ucFabV, but not the mutant, functionally replaced FabI within living cells. Conclusion: ucFabV confers increased tolerance through its function as an enoyl-ACP reductase. Furthermore, ucFabV is capable of restoring viability in the presence of compromised FabI, suggesting ucFabV is likely facilitating an alternate step within fatty acid synthesis, bypassing FabI inhibition.

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The effect of spatial structure on adaptation in Escherichia coli

Biology Letters ◽

10.1098/rsbl.2007.0481 ◽

2007 ◽

Vol 4 (1) ◽

pp. 57-59 ◽

Cited By ~ 33

Author(s):

Lilia Perfeito ◽

M. Inês Pereira ◽

Paulo R.A Campos ◽

Isabel Gordo

Keyword(s):

Escherichia Coli ◽

Spatial Structure ◽

Experimental Evolution ◽

Petri Dish ◽

Mutation Rates ◽

Wild Type ◽

Clonal Interference ◽

Genetic Studies ◽

Significant Difference ◽

The Cost

Populations of organisms are generally organized in a given spatial structure. However, the vast majority of population genetic studies are based on populations in which every individual competes globally. Here we use experimental evolution in Escherichia coli to directly test a recently made prediction that spatial structure slows down adaptation and that this cost increases with the mutation rate. This was studied by comparing populations of different mutation rates adapting to a liquid (unstructured) medium with populations that evolved in a Petri dish on solid (structured) medium. We find that mutators adapt faster to both environments and that adaptation is slower if there is spatial structure. We observed no significant difference in the cost of structure between mutator and wild-type populations, which suggests that clonal interference is intense in both genetic backgrounds.

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Antimutator Mutants in Bacteriophage T4 and Escherichia coli

Genetics ◽

10.1093/genetics/148.4.1579 ◽

1998 ◽

Vol 148 (4) ◽

pp. 1579-1585 ◽

Cited By ~ 1

Author(s):

Roel M Schaaper

Keyword(s):

Escherichia Coli ◽

Type Strain ◽

Wild Type Strain ◽

Bacteriophage T4 ◽

Mutation Rates ◽

Spontaneous Mutations ◽

Wild Type ◽

Mutant Strains ◽

Distinguishing Property ◽

Insight Into

Abstract Antimutators are mutant strains that have reduced mutation rates compared to the corresponding wild-type strain. Their existence, along with mutator mutants that have higher mutation rates compared to the wild-type strain, are powerful evidence that mutation rates are genetically controlled. Compared to mutator mutants, antimutators have a very distinguishing property. Because they prevent normally occurring mutations, they, uniquely, are capable of providing insight into the mechanisms of spontaneous mutations. In this review, antimutator mutants are discussed in bacteriophage T4 and the bacterium Escherichia coli, with regard to their properties, possible mechanisms, and implications for the sources of spontaneous mutations in these two organisms.

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Human mismatch-repair protein MutL homologue 1 (MLH1) interacts with Escherichia coli MutL and MutS in vivo and in vitro: a simple genetic system to assay MLH1 function

Biochemical Journal ◽

10.1042/bj20021205 ◽

2003 ◽

Vol 371 (1) ◽

pp. 183-189 ◽

Cited By ~ 8

Author(s):

Barbara QUARESIMA ◽

Pietro ALIFANO ◽

Pierfrancesco TASSONE ◽

Enrico V. AVVEDIMENTO ◽

Francesco S. COSTANZO ◽

...

Keyword(s):

Escherichia Coli ◽

Mismatch Repair ◽

Fold Increase ◽

Genetic System ◽

Mutation Rates ◽

Wild Type ◽

Mismatched Dna ◽

Repair Protein

A simple genetic system has been developed to test the effect of over-expression of wild-type or mutated human MutL homologue 1 (hMLH1) proteins on methyl-directed mismatch repair (MMR) in Escherichia coli. The system relies on detection of Lac+ revertants using MMR-proficient or MMR-deficient E. coli strains carrying a lac +1 frameshift mutation expressing hMLH1 proteins. We report that expression of wild-type hMLH1 protein causes an approx. 19-fold increase in mutation rates. The mutator phenotype was due to the ability of hMLH1 protein to interact with bacterial MutL and MutS proteins, thereby interfering with the formation of complexes between MMR proteins and mismatched DNA. Conversely, expression of proteins encoded by alleles deriving from hereditary-non-polyposis-colon-cancer (HNPCC) families decreases mutation rates, depending on the specific amino acid substitutions. These effects parallel the MutL-and MutS-binding and ATP-binding/hydrolysis activities of the mutated proteins.

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Mutation accumulation and fitness in mutator subpopulations of Escherichia coli

Biology Letters ◽

10.1098/rsbl.2012.0961 ◽

2013 ◽

Vol 9 (1) ◽

pp. 20120961 ◽

Cited By ~ 10

Author(s):

Ram P. Maharjan ◽

Bin Liu ◽

Yang Li ◽

Peter R. Reeves ◽

Lei Wang ◽

...

Keyword(s):

Escherichia Coli ◽

Significant Proportion ◽

Mutation Accumulation ◽

Mutation Rates ◽

Deleterious Mutations ◽

Wild Type ◽

Beneficial Mutations ◽

Bacterial Populations ◽

Neutral Mutations ◽

Over Time

Bacterial populations in clinical and laboratory settings contain a significant proportion of mutants with elevated mutation rates (mutators). Mutators have a particular advantage when multiple beneficial mutations are needed for fitness, as in antibiotic resistance. Nevertheless, high mutation rates potentially lead to increasing numbers of deleterious mutations and subsequently to the decreased fitness of mutators. To test how fitness changed with mutation accumulation, genome sequencing and fitness assays of nine Escherichia coli mutY mutators were undertaken in an evolving chemostat population at three time points. Unexpectedly, the fitness in members of the mutator subpopulation became constant despite a growing number of mutations over time. To test if the accumulated mutations affected fitness, we replaced each of the known beneficial mutations with wild-type alleles in a mutator isolate. We found that the other 25 accumulated mutations were not deleterious. Our results suggest that isolates with deleterious mutations are eliminated by competition in a continuous culture, leaving mutators with mostly neutral mutations. Interestingly, the mutator–non-mutator balance in the population reversed after the fitness plateau of mutators was reached, suggesting that the mutator–non-mutator ratio in populations has more to do with competition between members of the population than the accumulation of deleterious mutations.

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