Estimates of the Rate and Distribution of Fitness Effects of Spontaneous Mutation in Saccharomyces cerevisiae

Genetics ◽  
2001 ◽  
Vol 157 (1) ◽  
pp. 53-61 ◽  
Author(s):  
Clifford Zeyl ◽  
J Arjan G M DeVisser
Keyword(s):  
Mismatch Repair ◽  
Spontaneous Mutation ◽  
Evolutionary Genetics ◽  
Maximum Likelihood Estimates ◽  
Model Organisms ◽  
Wild Type ◽  
Mutator Strain ◽  
Slightly Deleterious Mutations ◽  
Mean Fitness ◽  
Change In Mean

Abstract The per-genome, per-generation rate of spontaneous mutation affecting fitness (U) and the mean fitness cost per mutation (s) are important parameters in evolutionary genetics, but have been estimated for few species. We estimated U and sh (the heterozygous effect of mutations) for two diploid yeast strains differing only in the DNA mismatch-repair deficiency used to elevate the mutation rate in one (mutator) strain. Mutations were allowed to accumulate in 50 replicate lines of each strain, during 36 transfers of randomly chosen single colonies (∼600 generations). Among wild-type lines, fitnesses were bimodal, with one mode showing no change in mean fitness. The other mode showed a mean 29.6% fitness decline and the petite phenotype, usually caused by partial deletion of the mitochondrial genome. Excluding petites, maximum-likelihood estimates adjusted for the effect of selection were U = 9.5 × 10-5 and sh = 0.217 for the wild type. Among the mutator lines, the best fit was obtained with 0.005 ≤ U ≤ 0.94 and 0.049 ≥ sh ≥ 0.0003. Like other recently tested model organisms, wild-type yeast have low mutation rates, with high mean fitness costs per mutation. Inactivation of mismatch repair increases the frequency of slightly deleterious mutations by approximately two orders of magnitude.

scholarly journals Functional Analysis of the NucS/EndoMS of the Hyperthermophilic Archaeon Sulfolobus islandicus REY15A

2020 ◽  
Vol 11 ◽  
Author(s):  
Sohail Ahmad ◽  
Qihong Huang ◽  
Jinfeng Ni ◽  
Yuanxi Xiao ◽  
Yunfeng Yang ◽  
...  
Keyword(s):  
Mismatch Repair ◽  
Mutation Rate ◽  
Spontaneous Mutation ◽  
Repair Pathway ◽  
Wild Type ◽  
Spontaneous Mutation Rate ◽  
Hyperthermophilic Archaeon ◽  
Cleavage Activity ◽  
Sulfolobus Islandicus ◽  
Mismatch Repair Pathway

EndoMS is a recently identified mismatch specific endonuclease in Thermococcales of Archaea and Mycobacteria of Bacteria. The homologs of EndoMS are conserved in Archaea and Actinobacteria, where classic MutS-MutL-mediated DNA mismatch repair pathway is absent or non-functional. Here, we report a study on the in vitro mismatch cleavage activity and in vivo function of an EndoMS homolog (SisEndoMS) from Sulfolobus islandicus REY15A, the model archaeon belonging to Crenarchaeota. SisEndoMS is highly active on duplex DNA containing G/T, G/G, and T/T mismatches. Interestingly, the cleavage activity of SisEndoMS is stimulated by the heterotrimeric PCNAs, and when Mn2+ was used as the co-factor instead of Mg2+, SisEndoMS was also active on DNA substrates containing C/T or A/G mismatches, suggesting that the endonuclease activity can be regulated by ion co-factors and accessory proteins. We compared the spontaneous mutation rate of the wild type strain REY15A and ∆endoMS by counter selection against 5-fluoroorotic acid (5-FOA). The endoMS knockout mutant had much higher spontaneous mutation rate (5.06 × 10−3) than that of the wild type (4.6 × 10−6). A mutation accumulation analysis also showed that the deletion mutant had a higher mutation occurrence than the wild type, with transition mutation being the dominant, suggesting that SisEndoMS is responsible for mutation avoidance in this hyperthermophilic archaeon. Overexpression of the wild type SisEndoMS in S. islandicus resulted in retarded growth and abnormal cell morphology, similar to strains overexpressing Hje and Hjc, the Holliday junction endonucleases. Transcriptomic analysis revealed that SisEndoMS overexpression led to upregulation of distinct gene including the CRISPR-Cas IIIB system, methyltransferases, and glycosyltransferases, which are mainly localized to specific regions in the chromosome. Collectively, our results support that EndoMS proteins represent a noncanonical DNA repair pathway in Archaea. The mechanism of the mismatch repair pathway in Sulfolobus which have a single chromosome is discussed.

scholarly journals The Consequences of Growth of a Mutator Strain of Escherichia coli as Measured by Loss of Function Among Multiple Gene Targets and Loss of Fitness

Genetics ◽  
2000 ◽  
Vol 154 (3) ◽  
pp. 959-970 ◽  
Author(s):  
Pauline Funchain ◽  
Annie Yeung ◽  
Jean Lee Stewart ◽  
Rose Lin ◽  
Malgorzata M Slupska ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Mismatch Repair ◽  
Growth Curves ◽  
Temperature Sensitive ◽  
Loss Of Function ◽  
Wild Type ◽  
Average Cell ◽  
Mutator Strain ◽  
Single Colony ◽  
Intergenic Regions

Abstract We have examined the composition of members of mutator populations of Escherichia coli by employing an extensive set of phenotypic screens that allow us to monitor the function of >700 genes, constituting ~15% of the genome. We looked at mismatch repair deficient cells after repeated cycles of single colony isolation on rich medium to generate lineages that are forced through severe bottlenecks, and compared the results to those for wild-type strains. The mutator lineages continued to accumulate mutations rapidly with each increasing cycle of colony isolation. By the end of the 40th cycle, after ~1000 generations, most of the lineages had reduced colony size, 4% had died out, 55% had auxotrophic requirements (increasing to 80% after 60 cycles), and 70% had defects in at least one sugar or catabolic pathway. In addition, 33% had a defect in cell motility, and 26% were either temperature-sensitive or cold-sensitive lethals. On the other hand, only 3% of the wild-type lineages had detectable mutations of any type after 40 cycles. By the 60th cycle, the typical mutator cell carried 4–5 inactive genes among the 15% of the genome being monitored, indicating that the average cell carried at least 24–30 inactivated genes distributed throughout the genome. Remarkably, 30% of the lineages had lost the ability to utilize xylose as a carbon source. DNA sequencing revealed that most of the Xyl− mutants had a frameshift in a run of eight G's (GGGGGGGG) in the xylB gene, either adding or deleting one -G-. Further analysis indicated that rendering E. coli deficient in mismatch repair unmasks hypermutable sites in certain genes or intergenic regions. Growth curves and competition tests on lineages that passed through 90 cycles of single colony isolation showed that all lineages suffered reduced fitness. We discuss these results in terms of the value of mutators in cellular evolution.

scholarly journals Escherichia coli mutator mutD5 is defective in the mutHLS pathway of DNA mismatch repair.

Genetics ◽  
1989 ◽  
Vol 121 (2) ◽  
pp. 205-212 ◽  
Author(s):  
R M Schaaper
Keyword(s):  
Escherichia Coli ◽  
Mismatch Repair ◽  
Patch Repair ◽  
Wild Type ◽  
Mutator Strain ◽  
Dna Mismatch ◽  
Repair Deficiency ◽  
In The Wild ◽  
Mutator Strains ◽  
Repair Defect

Abstract We have previously reported that the Escherichia coli mutator strain mutD5 was defective in the correction of bacteriophage M13mp2 heteroduplex DNA containing a T.G mismatch. Here, this defect was further investigated with regard to its interaction with the mutHLS pathway of mismatch repair. A set of 15 different M13mp2 heteroduplexes was used to measure the mismatch-repair capability of wild-type, mutL and mutD5 cells. Throughout the series, the mutD5 strain proved as deficient in mismatch repair as the mutL strain, indicating that the repair defect is similar in the two strains in both extent and specificity. [One exception was noted in the case a T.G mispair that was subject to VSP (Very Short Patch) repair. VSP repair was abolished by mutL but not by mutD.] Variation in the dam-methylation state of the heteroduplex molecules clearly affected repair in the wild-type strain but had no effect on either the mutD or mutL strain. Finally, mutDmutL or mutDmutS double-mutator strains were no more deficient in mismatch repair as were the single mutator strains. The combined results strongly argue that the mismatch-repair deficiency of mutD5 cells resides in the mutH,L,S-dependent pathway of mismatch repair and that the high mutation rate of mutD strains derives in part from this defect.

scholarly journals Spontaneous mutation at the mtr locus in neurospora: the molecular spectrum in wild-type and a mutator strain.

Genetics ◽  
1994 ◽  
Vol 138 (1) ◽  
pp. 61-74
Author(s):  
D Dillon ◽  
D Stadler
Keyword(s):  
Sequence Analysis ◽  
Spontaneous Mutation ◽  
Molecular Spectrum ◽  
Open Reading Frame ◽  
Wild Type ◽  
Spontaneous Mutation Rate ◽  
Reading Frame ◽  
Mutator Strain ◽  
Oak Ridge ◽  
Base Substitutions

Abstract Sequence analysis of 34 mtr mutations has yielded the first molecular spectrum of spontaneous mutants in Neurospora crassa. The great majority of the mutations are base substitutions (48%) or deletions (35%). In addition, sequence analysis of the entire mtr region, including the 1472-base pair open reading frame and 1205 base pairs of flanking DNA, was performed in both the Oak Ridge and Mauriceville strains of Neurospora, which are known to be divergent at the DNA level. Sixteen sequence differences between these two strains have been found in the mtr region, with 13 of these in DNA flanking the open reading frame. The differences consisted of base substitutions and small frameshifts at monotonic runs. This set of sequence differences has allowed a comparison of mutations in unselected DNA to those mutations that produce a phenotypic signal. We have isolated a mutator strain (mut-1) of Neurospora in which the spontaneous mutation rate at various loci is as much as 80-fold higher than in the non-mutator (wild type). Twenty-one mtr mutations in the mutator background have been sequenced and compared to the non-mutator spectrum, revealing a striking increase in -1 frameshift mutations. These frameshifts occur exclusively within or adjacent to monotonic runs and can be explained by small slippage events during DNA replication. This argues for a role of the mut-1 gene in this process.

The Budding Yeast Msh4 Protein Functions in Chromosome Synapsis and the Regulation of Crossover Distribution

Genetics ◽  
2001 ◽  
Vol 158 (3) ◽  
pp. 1013-1025 ◽  
Author(s):  
Janet E Novak ◽  
Petra B Ross-Macdonald ◽  
G Shirleen Roeder
Keyword(s):  
Mismatch Repair ◽  
Budding Yeast ◽  
Null Mutation ◽  
Crossing Over ◽  
Wild Type ◽  
Crossover Interference ◽  
Meiotic Chromosomes ◽  
Chromosome Synapsis ◽  
Protein Functions ◽  
Or Gene

AbstractThe budding yeast MSH4 gene encodes a MutS homolog produced specifically in meiotic cells. Msh4 is not required for meiotic mismatch repair or gene conversion, but it is required for wild-type levels of crossing over. Here, we show that a msh4 null mutation substantially decreases crossover interference. With respect to the defect in interference and the level of crossing over, msh4 is similar to the zip1 mutant, which lacks a structural component of the synaptonemal complex (SC). Furthermore, epistasis tests indicate that msh4 and zip1 affect the same subset of meiotic crossovers. In the msh4 mutant, SC formation is delayed compared to wild type, and full synapsis is achieved in only about half of all nuclei. The simultaneous defects in synapsis and interference observed in msh4 (and also zip1 and ndj1/tam1) suggest a role for the SC in mediating interference. The Msh4 protein localizes to discrete foci on meiotic chromosomes and colocalizes with Zip2, a protein involved in the initiation of chromosome synapsis. Both Zip2 and Zip1 are required for the normal localization of Msh4 to chromosomes, raising the possibility that the zip1 and zip2 defects in crossing over are indirect, resulting from the failure to localize Msh4 properly.

A Mutation in a Saccharomyces cerevisiae Gene (RAD3) Required for Nucleotide Excision Repair and Transcription Increases the Efficiency of Mismatch Correction

Genetics ◽  
1996 ◽  
Vol 144 (2) ◽  
pp. 459-466 ◽  
Author(s):  
Yingying Yang ◽  
Anthony L Johnson ◽  
Leland H Johnston ◽  
Wolfram Siede ◽  
Errol C Friedberg ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mismatch Repair ◽  
Excision Repair ◽  
Spontaneous Mutation ◽  
Surprising Result ◽  
Spontaneous Mutation Rate ◽  
Mismatch Correction ◽  
Nucleotide Excision ◽  
Repair Genes ◽  
Type Strains

Abstract RAD3 functions in DNA repair and transcription in Saccharomyces cerevisiae and particular rad3 alleles confer a mutator phenotype, possibly as a consequence of defective mismatch correction. We assessed the potential involvement of the Rad3 protein in mismatch correction by comparing heteroduplex repair in isogenic rad3-1 and wild-type strains. The rad3-1 allele increased the spontaneous mutation rate but did not prevent heteroduplex repair or bias its directionality. Instead, the efficiency of mismatch correction was enhanced in the rad3-1 strain. This surprising result prompted us to examine expression of yeast mismatch repair genes. We determined that MSH2, but not MLH1, is transcriptionally regulated during the cell-cycle like PMSl, and that rad3-1 does not increase the transcript levels for these genes in log phase cells. These observations suggest that the rad3-1 mutation gives rise to an enhanced efficiency of mismatch correction via a process that does not involve transcriptional regulation of mismatch repair. Interestingly, mismatch repair also was more efficient when error-editing by yeast DNA polymerase δ was eliminated. We discuss our results in relation to possible mechanisms that may link the rad3-1 mutation to mismatch correction efficiency.

scholarly journals Saturation of DNA Mismatch Repair and Error Catastrophe by a Base Analogue in Escherichia coli

Genetics ◽  
2002 ◽  
Vol 161 (4) ◽  
pp. 1363-1371
Author(s):  
Kazuo Negishi ◽  
David Loakes ◽  
Roel M Schaaper
Keyword(s):  
Escherichia Coli ◽  
Mismatch Repair ◽  
Dna Mismatch Repair ◽  
Repair System ◽  
Mutagenic Action ◽  
Wild Type ◽  
Dna Mismatch ◽  
Cell Counts ◽  
Error Catastrophe ◽  
Mismatch Repair System

Abstract Deoxyribosyl-dihydropyrimido[4,5-c][1,2]oxazin-7-one (dP) is a potent mutagenic deoxycytidine-derived base analogue capable of pairing with both A and G, thereby causing G · C → A · T and A · T → G · C transition mutations. We have found that the Escherichia coli DNA mismatch-repair system can protect cells against this mutagenic action. At a low dose, dP is much more mutagenic in mismatch-repair-defective mutH, mutL, and mutS strains than in a wild-type strain. At higher doses, the difference between the wild-type and the mutator strains becomes small, indicative of saturation of mismatch repair. Introduction of a plasmid containing the E. coli mutL+ gene significantly reduces dP-induced mutagenesis. Together, the results indicate that the mismatch-repair system can remove dP-induced replication errors, but that its capacity to remove dP-containing mismatches can readily be saturated. When cells are cultured at high dP concentration, mutant frequencies reach exceptionally high levels and viable cell counts are reduced. The observations are consistent with a hypothesis in which dP-induced cell killing and growth impairment result from excess mutations (error catastrophe), as previously observed spontaneously in proofreading-deficient mutD (dnaQ) strains.

scholarly journals Antagonism of Ultraviolet-Light Mutagenesis by the Methyl-Directed Mismatch-Repair System of Escherichia coli

Genetics ◽  
2000 ◽  
Vol 154 (2) ◽  
pp. 503-512 ◽  
Author(s):  
Hongbo Liu ◽  
Stephen R Hewitt ◽  
John B Hays
Keyword(s):  
Escherichia Coli ◽  
Mismatch Repair ◽  
Excision Repair ◽  
Spontaneous Mutation ◽  
Repair System ◽  
Uv Mutagenesis ◽  
Repair Protein ◽  
Mismatch Repair System

Abstract Previous studies have demonstrated that the Escherichia coli MutHLS mismatch-repair system can process UV-irradiated DNA in vivo and that the human MSH2·MSH6 mismatch-repair protein binds more strongly in vitro to photoproduct/base mismatches than to “matched” photoproducts in DNA. We tested the hypothesis that mismatch repair directed against incorrect bases opposite photoproducts might reduce UV mutagenesis, using two alleles at E. coli lacZ codon 461, which revert, respectively, via CCC → CTC and CTT → CTC transitions. F′ lacZ targets were mated from mut+ donors into mutH, mutL, or mutS recipients, once cells were at substantial densities, to minimize spontaneous mutation prior to irradiation. In umu+ mut+ recipients, a range of UV fluences induced lac+ revertant frequencies of 4–25 × 10−8; these frequencies were consistently 2-fold higher in mutH, mutL, or mutS recipients. Since this effect on mutation frequency was unaltered by an Mfd− defect, it appears not to involve transcription-coupled excision repair. In mut+ umuC122::Tn5 bacteria, UV mutagenesis (at 60 J/m2) was very low, but mutH or mutL or mutS mutations increased reversion of both lacZ alleles roughly 25-fold, to 5–10 × 10−8. Thus, at UV doses too low to induce SOS functions, such as Umu2′D, most incorrect bases opposite occasional photoproducts may be removed by mismatch repair, whereas in heavily irradiated (SOS-induced) cells, mismatch repair may only correct some photoproduct/base mismatches, so UV mutagenesis remains substantial.

scholarly journals Mutants of downy mildew resistance in Lactuca sativa (lettuce).

Genetics ◽  
1994 ◽  
Vol 137 (3) ◽  
pp. 867-874
Author(s):  
P A Okubara ◽  
P A Anderson ◽  
O E Ochoa ◽  
R W Michelmore
Keyword(s):  
Molecular Marker ◽  
Downy Mildew ◽  
Genomic Dna ◽  
Spontaneous Mutation ◽  
Multigene Families ◽  
Downy Mildew Resistance ◽  
Bremia Lactucae ◽  
Wild Type ◽  
Mildew Resistance ◽  
Recessive Mutations

Abstract As part of our investigation of disease resistance in lettuce, we generated mutants that have lost resistance to Bremia lactucae, the casual fungus of downy mildew. Using a rapid and reliable screen, we identified 16 distinct mutants of Latuca sativa that have lost activity of one of four different downy mildew resistance genes (Dm). In all mutants, only a single Dm specificity was affected. Genetic analysis indicated that the lesions segregated as single, recessive mutations at the Dm loci. Dm3 was inactivated in nine of the mutants. One of five Dm 1 mutants was selected from a population of untreated seeds and therefore carried a spontaneous mutation. All other Dm1, Dm3, Dm5/8 and Dm7 mutants were derived from gamma- or fast neutron-irradiated seed. In two separate Dm 1 mutants and in each of the eight Dm3 mutants analyzed, at least one closely linked molecular marker was absent. Also, high molecular weight genomic DNA fragments that hybridized to a tightly linked molecular marker in wild type were either missing entirely or were truncated in two of the Dm3 mutants, providing additional evidence that deletions had occurred in these mutants. Absence of mutations at loci epistatic to the Dm genes suggested that such loci were either members of multigene families, were critical for plant survival, or encoded components of duplicated pathways for resistance; alternatively, the genes determining downy mildew resistance might be limited to the Dm loci.

The Effect of Mismatch Repair and Heteroduplex Formation on Sexual Isolation in Bacillus

Genetics ◽  
1998 ◽  
Vol 148 (1) ◽  
pp. 13-18 ◽  
Author(s):  
Jacek Majewski ◽  
Frederick M Cohan
Keyword(s):  
Mismatch Repair ◽  
Wild Type Strain ◽  
Resistance Mechanism ◽  
Sequence Divergence ◽  
Sexual Isolation ◽  
Wild Type ◽  
Exponential Relationship ◽  
Dna Strands ◽  
The Relationship ◽  
Heteroduplex Formation

AbstractIn Bacillus transformation, sexual isolation is known to be an exponential function of the sequence divergence between donor and recipient. Here, we have investigated the mechanism under which sequence divergence results in sexual isolation. We tested the effect of mismatch repair by comparing a wild-type strain and an isogenic mismatch-repair mutant for the relationship between sexual isolation and sequence divergence. Mismatch repair was shown to contribute to sexual isolation but was responsible for only a small fraction of the sexual isolation observed. Another possible mechanism of sexual isolation is that more divergent recipient and donor DNA strands have greater difficulty forming a heteroduplex because a region of perfect identity between donor and recipient is required for initiation of the heteroduplex. A mathematical model showed that this heteroduplex-resistance mechanism yields an exponential relationship between sexual isolation and sequence divergence. Moreover, this model yields an estimate of the size of the region of perfect identity that is comparable to independent estimates for Escherichia coli. For these reasons, and because all other mechanisms of sexual isolation may be ruled out, we conclude that resistance to heteroduplex formation is predominantly responsible for the exponential relationship between sexual isolation and sequence divergence in Bacillus transformation.

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