scholarly journals Reduced Mismatch Repair of Heteroduplexes Reveals “Non”-interfering Crossing Over in Wild-Type Saccharomyces cerevisiae

Genetics ◽  
2008 ◽  
Vol 178 (3) ◽  
pp. 1251-1269 ◽  
Author(s):  
Tony J. Getz ◽  
Stephen A. Banse ◽  
Lisa S. Young ◽  
Allison V. Banse ◽  
Johanna Swanson ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mismatch Repair ◽  
Crossing Over ◽  
Wild Type

scholarly journals Analysis of Conditional Mutations in the Saccharomyces cerevisiae MLH1 Gene in Mismatch Repair and in Meiotic Crossing Over

Genetics ◽  
2002 ◽  
Vol 160 (3) ◽  
pp. 909-921 ◽  
Author(s):  
Juan Lucas Argueso ◽  
Daniel Smith ◽  
James Yi ◽  
Marc Waase ◽  
Sumeet Sarin ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Family ◽  
Mismatch Repair ◽  
Mutational Analysis ◽  
Mutation Rates ◽  
Temperature Sensitive ◽  
Crossing Over ◽  
Wild Type ◽  
Genetic Studies ◽  
Conditional Mutations

Abstract In mismatch repair (MMR), members of the MLH gene family have been proposed to act as key molecular matchmakers to coordinate mismatch recognition with downstream repair functions that result in mispair excision. Two members of this gene family, MLH1 and MLH3, have also been implicated in meiotic crossing over. These diverse roles suggest that a mutational analysis of MLH genes could provide reagents required to identify interactions between gene products and to test whether the different roles ascribed to a subset of these genes can be separated. In this report we show that in Saccharomyces cerevisiae the mlh1Δ mutation confers inviability in pol3-01 strain backgrounds that are defective in the Polδ proofreading exonuclease activity. This phenotype was exploited to identify four mlh1 alleles that each confer a temperature-sensitive phenotype for viability in pol3-01 strains. In three different mutator assays, strains bearing conditional mlh1 alleles displayed wild-type or nearly wild-type mutation rates at 26°. At 35°, these strains exhibited mutation rates that approached those observed in mlh1Δ mutants. The mutator phenotype exhibited in mlh1-I296S strains was partially suppressed at 35° by EXO1 overexpression. The mlh1-F228S and -I296S mutations conferred a separation-of-function phenotype in meiosis; both mlh1-F228S and -I296S strains displayed strong defects in meiotic mismatch repair but showed nearly wild-type levels of crossing over, suggesting that the conditional mutations differentially affected MLH1 functions. These genetic studies suggest that the conditional mlh1 mutations can be used to separate the MMR and meiotic crossing-over functions of MLH1 and to identify interactions between MLH1 and downstream repair components.

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.

scholarly journals The Conversion Gradient at HIS4 of Saccharomyces cerevisiae. II. A Role for Mismatch Repair Directed by Biased Resolution of the Recombinational Intermediate

Genetics ◽  
1999 ◽  
Vol 153 (2) ◽  
pp. 573-583 ◽  
Author(s):  
Henriette M Foss ◽  
Kenneth J Hillers ◽  
Franklin W Stahl
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Synthesis ◽  
Mismatch Repair ◽  
Strand Break ◽  
Double Strand Break ◽  
Double Strand Break Repair ◽  
Holliday Junction ◽  
Crossing Over ◽  
Gradient Type ◽  
Salient Features

AbstractSalient features of recombination at ARG4 of Saccharomyces provoke a variation of the double-strand-break repair (DSBR) model that has the following features: (1) Holliday junction cutting is biased in favor of strands upon which DNA synthesis occurred during formation of the joint molecule (this bias ensures that cutting both junctions of the joint-molecule intermediate arising during DSBR usually leads to crossing over); (2) cutting only one junction gives noncrossovers; and (3) repair of mismatches that are semirefractory to mismatch repair and/or far from the DSB site is directed primarily by junction resolution. The bias in junction resolution favors restoration of 4:4 segregation when such mismatches and the directing junction are on the same side of the DSB site. Studies at HIS4 confirmed the predicted influence of the bias in junction resolution on the conversion gradient, type of mismatch repair, and frequency of aberrant 5:3 segregation, as well as the predicted relationship between mismatch repair and crossing over.

scholarly journals Systematic Mutagenesis of the Saccharomyces cerevisiae MLH1 Gene Reveals Distinct Roles for Mlh1p in Meiotic Crossing Over and in Vegetative and Meiotic Mismatch Repair

2003 ◽  
Vol 23 (3) ◽  
pp. 873-886 ◽  
Author(s):  
Juan Lucas Argueso ◽  
Amanda Wraith Kijas ◽  
Sumeet Sarin ◽  
Julie Heck ◽  
Marc Waase ◽  
...  
Keyword(s):  
Mismatch Repair ◽  
Ternary Complex ◽  
Genetic Recombination ◽  
Crossing Over ◽  
Wild Type ◽  
Dna Mismatch ◽  
Biochemical Assays ◽  
Postmeiotic Segregation ◽  
New Alleles

ABSTRACT In eukaryotic cells, DNA mismatch repair is initiated by a conserved family of MutS (Msh) and MutL (Mlh) homolog proteins. Mlh1 is unique among Mlh proteins because it is required in mismatch repair and for wild-type levels of crossing over during meiosis. In this study, 60 new alleles of MLH1 were examined for defects in vegetative and meiotic mismatch repair as well as in meiotic crossing over. Four alleles predicted to disrupt the Mlh1p ATPase activity conferred defects in all functions assayed. Three mutations, mlh1-2, -29, and -31, caused defects in mismatch repair during vegetative growth but allowed nearly wild-type levels of meiotic crossing over and spore viability. Surprisingly, these mutants did not accumulate high levels of postmeiotic segregation at the ARG4 recombination hotspot. In biochemical assays, Pms1p failed to copurify with mlh1-2, and two-hybrid studies indicated that this allele did not interact with Pms1p and Mlh3p but maintained wild-type interactions with Exo1p and Sgs1p. mlh1-29 and mlh1-31 did not alter the ability of Mlh1p-Pms1p to form a ternary complex with a mismatch substrate and Msh2p-Msh6p, suggesting that the region mutated in these alleles could be responsible for signaling events that take place after ternary complex formation. These results indicate that mismatches formed during genetic recombination are processed differently than during replication and that, compared to mismatch repair functions, the meiotic crossing-over role of MLH1 appears to be more resistant to mutagenesis, perhaps indicating a structural role for Mlh1p during crossing over.

Spontaneous Frameshift Mutations in Saccharomyces cerevisiae: Accumulation During DNA Replication and Removal by Proofreading and Mismatch Repair Activities

Genetics ◽  
2001 ◽  
Vol 159 (1) ◽  
pp. 65-75 ◽  
Author(s):  
Christopher N Greene ◽  
Sue Jinks-Robertson
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mismatch Repair ◽  
Wild Type Strain ◽  
Dna Polymerases ◽  
Wild Type ◽  
Triple Mutant ◽  
Repair Capacity ◽  
Frameshift Mutations ◽  
Dna Polymerization ◽  
In The Wild

Abstract The accumulation of frameshift mutations during DNA synthesis is determined by the rate at which frameshift intermediates are generated during DNA polymerization and the efficiency with which frameshift intermediates are removed by DNA polymerase-associated exonucleolytic proofreading activity and/or the postreplicative mismatch repair machinery. To examine the relative contributions of these factors to replication fidelity in Saccharomyces cerevisiae, we determined the reversion rates and spectra of the lys2ΔBgl +1 frameshift allele. Wild-type and homozygous mutant diploid strains with all possible combinations of defects in the exonuclease activities of DNA polymerases δ and ε (conferred by the pol3-01 and pol2-4 alleles, respectively) and in mismatch repair (deletion of MSH2) were analyzed. Although there was no direct correlation between homopolymer run length and frameshift accumulation in the wild-type strain, such a correlation was evident in the triple mutant strain lacking all repair capacity. Furthermore, examination of strains defective in one or two repair activities revealed distinct biases in the removal of the corresponding frameshift intermediates by exonucleolytic proofreading and/or mismatch repair. Finally, these analyses suggest that the mismatch repair machinery may be important for generating some classes of frameshift mutations in yeast.

scholarly journals MAPPING AND GENE CONVERSION STUDIES WITH THE STRUCTURAL GENE FOR ISO-1-CYTOCHROME C IN YEAST

Genetics ◽  
1975 ◽  
Vol 81 (4) ◽  
pp. 615-629
Author(s):  
Christopher W Lawrence ◽  
Fred Sherman ◽  
Mary Jackson ◽  
Richard A Gilmore
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Conversion ◽  
The Other ◽  
Crossing Over ◽  
Wild Type ◽  
Chromosome X ◽  
Tetrad Analysis ◽  
Yeast Saccharomyces Cerevisiae ◽  
Distal Marker ◽  
The Right

ABSTRACT We have investigated the order of the four genes cyc1, rad7, SUP4, and cdc8 which form a tightly linked cluster on the right arm of chromosome X in the yeast Saccharomyces cerevisiae. Crossing over and coconversion data from tetrad analysis established the gene order to be centromere–cyc1–rad7–SUP4. Also cdc8 appeared to be distal to SUP4 on the basis of crossovers that were associated with conversion of SUP4. The frequencies of recombination and the occurrence of coconversions suggest that these four genes are contiguous or at least nearly so. Gene-conversion frequencies for several cyc1 alleles were studied, including cyc1–1, a deletion of the whole gene that extends into the rad7 locus. The cyc1–1 deletion was found to be capable of conversion, though at a frequency some fivefold less than the other alleles studied, and both 3:1 and 1:3 events were detected. In general 1:3 and 3:1 conversion events were equally frequent at all loci studied, and approximately 50% of conversions were accompanied by reciprocal recombination for flanking markers. The orientation of the cyc1 gene could not be clearly deduced from the behavior of the distal marker SUP4 in wild-type recombinants that arose from diploids heteroallelic for cyc1 mutations.

scholarly journals Meiotic mismatch repair quantified on the basis of segregation patterns in Schizosaccharomyces pombe.

Genetics ◽  
1993 ◽  
Vol 133 (4) ◽  
pp. 815-824 ◽  
Author(s):  
P Schär ◽  
P Munz ◽  
J Kohli
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Schizosaccharomyces Pombe ◽  
Detailed Analysis ◽  
Mismatch Repair ◽  
Meiotic Recombination ◽  
Wild Type ◽  
Base Pairs ◽  
Ascobolus Immersus ◽  
Postmeiotic Segregation ◽  
Hybrid Dna

Abstract Hybrid DNA with mismatched base pairs is a central intermediate of meiotic recombination. Mismatch repair leads either to restoration or conversion, while failure of repair results in postmeiotic segregation (PMS). The behavior of three G to C transversions in one-factor crosses with the wild-type alleles is studied in Schizosaccharomyces pombe. They lead to C/C and G/G mismatches and are compared with closely linked mutations yielding other mismatches. A method is presented for the detection of PMS in random spores. The procedure yields accurate PMS frequencies as shown by comparison with tetrad data. A scheme is presented for the calculation of the frequency of hybrid DNA formation and the efficiency of mismatch repair. The efficiency of C/C repair in S. pombe is calculated to be about 70%. Other mismatches are repaired with close to 100% efficiency. These results are compared with data published on mutations in Saccharomyces cerevisiae and Ascobolus immersus. This study forms the basis for the detailed analysis of the marker effects caused by G to C transversions in two-factor crosses.

scholarly journals A Saccharomyces cerevisiae RAD52 allele expressing a C-terminal truncation protein: activities and intragenic complementation of missense mutations.

Genetics ◽  
1993 ◽  
Vol 133 (1) ◽  
pp. 39-49 ◽  
Author(s):  
K L Boundy-Mills ◽  
D M Livingston
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Repair ◽  
Deletion Mutants ◽  
Crossing Over ◽  
Missense Mutations ◽  
Wild Type ◽  
Spore Viability ◽  
Deletion Allele ◽  
Low Level ◽  
Intragenic Complementation

Abstract A nonsense allele of the yeast RAD52 gene, rad52-327, which expresses the N-terminal 65% of the protein was compared to two missense alleles, rad52-1 and rad52-2, and to a deletion allele. While the rad52-1 and the deletion mutants have severe defects in DNA repair, recombination and sporulation, the rad52-327 and rad52-2 mutants retain either partial or complete capabilities in repair and recombination. These two mutants behave similarly in most tests of repair and recombination during mitotic growth. One difference between these two alleles is that a homozygous rad52-2 diploid fails to sporulate, whereas the homozygous rad52-327 diploid sporulates weakly. The low level of sporulation by the rad52-327 diploid is accompanied by a low percentage of spore viability. Among these viable spores the frequency of crossing over for markers along chromosome VII is the same as that found in wild-type spores. rad52-327 complements rad52-2 for repair and sporulation. Weaker intragenic complementation occurs between rad52-327 and rad52-1.

scholarly journals Saccharomyces cerevisiae Mer3 Is a DNA Helicase Involved in Meiotic Crossing Over

2002 ◽  
Vol 22 (10) ◽  
pp. 3281-3291 ◽  
Author(s):  
Takuro Nakagawa ◽  
Richard D. Kolodner
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Helicase ◽  
Double Strand Breaks ◽  
Crossing Over ◽  
Helicase Activity ◽  
Wild Type ◽  
Rna Helicases ◽  
Conserved Residues ◽  
Strand Breaks ◽  
Homologous Chromosomes

ABSTRACT Crossing over is regulated to occur at least once per each pair of homologous chromosomes during meiotic prophase to ensure proper segregation of chromosomes at the first meiotic division. In a mer3 deletion mutant of Saccharomyces cerevisiae, crossing over is decreased, and the distribution of the crossovers that occur is random. The predicted Mer3 protein contains seven motifs characteristic of the DExH box type of DNA/RNA helicases. The mer3G166D and the mer3K167A mutation, amino acid substitutions of conserved residues in a putative nucleotide-binding domain of the helicase motifs caused a defect in the transition of meiosis-specific double-strand breaks to later intermediates, decreased crossing over, and reduced crossover interference. The purified Mer3 protein was found to have DNA helicase activity. This helicase activity was reduced by the mer3GD mutation to <1% of the wild-type activity, even though binding of the mutant protein to single- and double-strand DNA was unaffected. The mer3KA mutation eliminated the ATPase activity of the wild-type protein. These results demonstrate that Mer3 is a DNA helicase that functions in meiotic crossing over.

Control of meiotic recombination in Arabidopsis: role of the MutL and MutS homologues

2006 ◽  
Vol 34 (4) ◽  
pp. 542-544 ◽  
Author(s):  
F.C.H. Franklin ◽  
J.D. Higgins ◽  
E. Sanchez-Moran ◽  
S.J. Armstrong ◽  
K.E. Osman ◽  
...  
Keyword(s):  
Mismatch Repair ◽  
Meiotic Recombination ◽  
Holliday Junction ◽  
Crossing Over ◽  
Wild Type ◽  
Pathway Model ◽  
Junction Formation ◽  
Double Holliday Junction ◽  
Dependent Pathway

Immunocytochemistry reveals that the Arabidopsis mismatch repair proteins AtMSH4, AtMLH3 and AtMLH1 are expressed during prophase I of meiosis. Expression of AtMSH4 precedes AtMLH3 and AtMLH1 which co-localize as foci during pachytene. Co-localization between AtMSH4 and AtMLH3 occurs, but appears transient. AtMLH3 foci are not detected in an Atmsh4 mutant. However, localization of AtMSH4 is unaffected in Atmlh3, suggesting that recombination may proceed to dHj (double Holliday junction) formation. Mean chiasma frequency in Atmsh4 is reduced to 1.55 compared with 9.86 in wild-type. In contrast with wild-type, the distribution of residual crossovers in Atmsh4 closely fits a Poisson distribution. This is consistent with a two-pathway model for meiotic crossing-over whereby most crossovers occur via an AtMSH4-dependent pathway that is subject to interference, with the remaining crossovers arising via an interference-independent pathway. Loss of AtMLH3 results in an approx. 60% reduction in crossovers. Results suggest that dHj resolution can occur, but in contrast with wild-type where most or all dHjs are directed to form crossovers, the outcome is biased in favour of a non-crossover outcome. The results are compatible with a model whereby the MutL complex maintains or imposes a dHj conformation that ensures crossover formation.

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