Substrate length requirements for efficient mitotic recombination in Saccharomyces cerevisiae

1993 ◽  
Vol 13 (7) ◽  
pp. 3937-3950
Author(s):  
S Jinks-Robertson ◽  
M Michelitch ◽  
S Ramcharan
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Conversion ◽  
Mitotic Recombination ◽  
Yeast Genome ◽  
Inverted Repeats ◽  
Ectopic Recombination ◽  
Linear Relationships ◽  
Recombination System ◽  
Efficient Processing ◽  
Substrate Size

An ectopic recombination system using ura3 heteroalleles varying in size from 80 to 960 bp has been used to examine the effect of substrate length on spontaneous mitotic recombination. The ura3 heteroalleles were positioned either on nonhomologous chromosomes (heterochromosomal repeats) or as direct or inverted repeats on the same chromosome (intrachromosomal repeats). While the intrachromosomal events occur at rates at least 2 orders of magnitude greater than the corresponding heterochromosomal events, the recombination rate for each type of repeat considered separately exhibits a linear dependence on substrate length. The linear relationships allow estimation of the corresponding minimal efficient processing segments, which are approximately 250 bp regardless of the relative positions of the repeats in the yeast genome. An examination of the distribution of recombination events into simple gene conversion versus crossover events indicates that reciprocal exchange is more sensitive to substrate size than is gene conversion.

scholarly journals Substrate length requirements for efficient mitotic recombination in Saccharomyces cerevisiae.

1993 ◽  
Vol 13 (7) ◽  
pp. 3937-3950 ◽  
Author(s):  
S Jinks-Robertson ◽  
M Michelitch ◽  
S Ramcharan
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Conversion ◽  
Mitotic Recombination ◽  
Yeast Genome ◽  
Inverted Repeats ◽  
Ectopic Recombination ◽  
Linear Relationships ◽  
Recombination System ◽  
Efficient Processing ◽  
Substrate Size

An ectopic recombination system using ura3 heteroalleles varying in size from 80 to 960 bp has been used to examine the effect of substrate length on spontaneous mitotic recombination. The ura3 heteroalleles were positioned either on nonhomologous chromosomes (heterochromosomal repeats) or as direct or inverted repeats on the same chromosome (intrachromosomal repeats). While the intrachromosomal events occur at rates at least 2 orders of magnitude greater than the corresponding heterochromosomal events, the recombination rate for each type of repeat considered separately exhibits a linear dependence on substrate length. The linear relationships allow estimation of the corresponding minimal efficient processing segments, which are approximately 250 bp regardless of the relative positions of the repeats in the yeast genome. An examination of the distribution of recombination events into simple gene conversion versus crossover events indicates that reciprocal exchange is more sensitive to substrate size than is gene conversion.

scholarly journals Characterization of the Roles of the Saccharomyces cerevisiae RAD54 Gene and a Homologue of RAD54, RDH54/TID1, in Mitosis and Meiosis

Genetics ◽  
1997 ◽  
Vol 147 (4) ◽  
pp. 1545-1556 ◽  
Author(s):  
Miki Shinohara ◽  
Emi Shita-Yamaguchi ◽  
Jean-Marie Buerstedde ◽  
Hideo Shinagawa ◽  
Hideyuki Ogawa ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Conversion ◽  
Double Mutant ◽  
Mitotic Recombination ◽  
Genome Project ◽  
Yeast Genome ◽  
Wild Type ◽  
Reduced Frequency ◽  
A Minor ◽  
Mitosis And Meiosis

Abstract The RAD54 gene, which encodes a protein in the SW12/SNF2 family, plays an important role in recombination and DNA repair in Saccharomyces cerevisiae. The yeast genome project revealed a homologue of RAD54, RDH54/TID1. Properties of the rdh54/tid1 mutant and the rad54 rdh54/tid1 double mutant are shown for mitosis and meiosis. The rad54 mutant is sensitive to the alkylating agent, methyl methanesulfonate (MMS), and is defective in interchromosomal and intrachromosomal gene conversion. The rdh54/tid1 single mutant, on the other hand, does not show any significant deficiency in mitosis. However, the rad54 rdh54/tid1 mutant is more sensitive to MMS and more defective in interchromosomal gene conversion than is the rad54 mutant, but shows the same frequency of intrachromosomal gene conversion as the rad54 mutant. These results suggest that RDH54/TID1 is involved in a minor pathway of mitotic recombination in the absence of RAD54. In meiosis, both single mutants produce viable spores at slightly reduced frequency. However, only the rdh54/tid1 mutant, but not the rad54 mutant, shows significant defects in recombination: retardation of the repair of meiosis-specific double-strand breaks (DSBs) and delayed formation of physical recombinants. Furthermore, the rad54 rdh54/tid1 double mutant is completely defective in meiosis, accumulating DSBs with more recessed ends than the wild type and producing fewer physical recombinants than the wild type. These results suggest that one of the differences between the late stages of mitotic recombination and meiotic recombination might be specified by differential dependency on the Rad54 and Rdh54/Tid1 proteins.

scholarly journals A defect in mismatch repair in Saccharomyces cerevisiae stimulates ectopic recombination between homeologous genes by an excision repair dependent process.

Genetics ◽  
1990 ◽  
Vol 126 (3) ◽  
pp. 535-547 ◽  
Author(s):  
A M Bailis ◽  
R Rothstein
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Repair ◽  
Gene Conversion ◽  
Mismatch Repair ◽  
Excision Repair ◽  
Mitotic Recombination ◽  
Mismatch Repair Gene ◽  
Repair Gene ◽  
Ectopic Recombination ◽  
Homeologous Genes

Abstract Null mutations in three recombination and DNA repair genes were studied to determine their effects on mitotic recombination between the duplicate AdoMet (S-adenosylmethionine) synthetase genes (SAM1 and SAM2) in Saccharomyces cerevisiae. SAM1 and SAM2, located on chromosomes XII and IV, respectively, encode functionally equivalent although differentially regulated AdoMet synthetases. These similar but not identical (homeologous) genes are 83% homologous at the nucleotide level and this identity is limited solely to the coding regions of the genes. Single frameshift mutations were introduced into the 5' end of SAM1 and the 3' end of SAM2 by restriction site ablation. The sequences surrounding these mutations differ significantly in their degree of homology to the corresponding area of the other gene. Mitotic ectopic recombination between the mutant sam genes occurs at a rate of 8.4 x 10(-9) in a wild-type genetic background. Gene conversion of the marker within the region of greater sequence homology occurs 20-fold more frequently than conversion of the marker within the region of relative sequence diversity. The relative orientation of the two genes prevents the recovery of translocations. Mitotic recombination between the sam genes is completely dependent on the DNA repair and recombination gene RAD52. A mutation in PMS1, a mismatch repair gene, causes a 4.5-fold increase in the rate of ectopic recombination. RAD1, an excision repair gene, is required to observe this increased rate of ectopic conversion. In addition, RAD1 is involved in modulating the pattern of coconversion during recombination between the homeologous sam genes. These results suggest that interactions between mismatch repair, excision repair and recombinational repair functions are involved in determining the ectopic gene conversion frequency between the sam genes.

scholarly journals Regulation of Mitotic Homeologous Recombination in Yeast: Functions of Mismatch Repair and Nucleotide Excision Repair Genes

Genetics ◽  
2000 ◽  
Vol 154 (1) ◽  
pp. 133-146 ◽  
Author(s):  
Ainsley Nicholson ◽  
Miyono Hendrix ◽  
Sue Jinks-Robertson ◽  
Gray F Crouse
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mismatch Repair ◽  
Nucleotide Excision Repair ◽  
Excision Repair ◽  
Mitotic Recombination ◽  
Inverted Repeats ◽  
Replication Errors ◽  
Nucleotide Excision ◽  
Homeologous Recombination ◽  
Repair Genes

Abstract The Saccharomyces cerevisiae homologs of the bacterial mismatch repair proteins MutS and MutL correct replication errors and prevent recombination between homeologous (nonidentical) sequences. Previously, we demonstrated that Msh2p, Msh3p, and Pms1p regulate recombination between 91% identical inverted repeats, and here use the same substrates to show that Mlh1p and Msh6p have important antirecombination roles. In addition, substrates containing defined types of mismatches (base-base mismatches; 1-, 4-, or 12-nt insertion/deletion loops; or 18-nt palindromes) were used to examine recognition of these mismatches in mitotic recombination intermediates. Msh2p was required for recognition of all types of mismatches, whereas Msh6p recognized only base-base mismatches and 1-nt insertion/deletion loops. Msh3p was involved in recognition of the palindrome and all loops, but also had an unexpected antirecombination role when the potential heteroduplex contained only base-base mismatches. In contrast to their similar antimutator roles, Pms1p consistently inhibited recombination to a lesser degree than did Msh2p. In addition to the yeast MutS and MutL homologs, the exonuclease Exo1p and the nucleotide excision repair proteins Rad1p and Rad10p were found to have roles in inhibiting recombination between mismatched substrates.

Effect of limited homology on gene conversion in a Saccharomyces cerevisiae plasmid recombination system

1988 ◽  
Vol 8 (6) ◽  
pp. 2442-2448 ◽  
Author(s):  
B Y Ahn ◽  
K J Dornfeld ◽  
T J Fagrelius ◽  
D M Livingston
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Conversion ◽  
Dna Sequences ◽  
Homologous Sequence ◽  
Insertion Mutation ◽  
Base Pairs ◽  
Recombination System ◽  
Plasmid Recombination ◽  
His3 Gene ◽  
Conversion Tract

Plasmids containing heteroallelic copies of the Saccharomyces cerevisiae HIS3 gene undergo intramolecular gene conversion in mitotically dividing S. cerevisiae cells. We have used this plasmid system to determine the minimum amount of homology required for gene conversion, to examine how conversion tract lengths are affected by limited homology, and to analyze the role of flanking DNA sequences on the pattern of exchange. Plasmids with homologous sequences greater than 2 kilobases have mitotic exchange rates as high as 2 x 10(-3) events per cell per generation. As the homology is reduced, the exchange rate decreases dramatically. A plasmid with 26 base pairs (bp) of homology undergoes gene conversion at a rate of approximately 1 x 10(-10) events per cell per generation. These studies have also shown that an 8-bp insertion mutation 13 bp from a border between homologous and nonhomologous sequences undergoes conversion, but that a similar 8-bp insertion 5 bp from a border does not. Examination of independent conversion events which occurred in plasmids with heteroallelic copies of the HIS3 gene shows that markers within 280 bp of a border between homologous and nonhomologous sequences undergo conversion less frequently than the same markers within a more extensive homologous sequence. Thus, proximity to a border between homologous and nonhomologous sequences shortens the conversion tract length.

RAD52-INDEPENDENT MITOTIC GENE CONVERSION IN SACCHAROMYCES CEREVISIAE FREQUENTLY RESULTS IN CHROMOSOMAL LOSS

Genetics ◽  
1985 ◽  
Vol 111 (1) ◽  
pp. 7-22
Author(s):  
James E Haber ◽  
Mark Hearn
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Conversion ◽  
Mitotic Recombination ◽  
Gene Conversion Event ◽  
Intragenic Recombination ◽  
Wild Type ◽  
Conversion Event ◽  
Chromosomal Loss ◽  
Mitotic Gene Conversion ◽  
Striking Effect

ABSTRACT We have examined spontaneous, interchromosomal mitotic recombination events between his4 alleles in both Rad+ and rad52 strains of Saccharomyces cerevisiae. In Rad+ strains, 74% of the His+ prototrophs resulted from gene conversion events without exchange of flanking markers. In diploids homozygous for the rad52-1 mutation, the frequency of His+ prototroph formation was less than 5% of the wild-type value, and more than 80% of the gene conversion events were accompanied by an exchange of flanking markers. Most of the rad52 intragenic recombination events arose by gene conversion accompanied by an exchange of flanking markers and not by a simple reciprocal exchange between the his4A and his4C alleles. There were also profound effects on the kinds of recombinant products that were recovered. The most striking effect was that RAD52-independent mitotic recombination frequently results in the loss of one of the two chromosomes participating in the gene conversion event.

Ectopic recombination between Ty elements in Saccharomyces cerevisiae is not induced by DNA damage

1992 ◽  
Vol 12 (10) ◽  
pp. 4441-4448
Author(s):  
A Parket ◽  
M Kupiec
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Damage ◽  
Uv Irradiation ◽  
Mitotic Recombination ◽  
Methyl Methanesulfonate ◽  
Ectopic Recombination ◽  
Ty Elements ◽  
Chemical Agents ◽  
Physical And Chemical

Mitotic recombination is increased when cells are treated with a variety of physical and chemical agents that cause damage to their DNA. We show here, using Saccharomyces cerevisiae strains that carry marked Ty elements, that recombination between members of this family of retrotransposons is not increased by UV irradiation or by treatment with the radiomimetic drug methyl methanesulfonate. Both ectopic recombination and mutation events were elevated by these agents for non-Ty sequences in the same strain. We discuss possible mechanisms that can prevent the induction of recombination between Ty elements.

scholarly journals Genetic Requirements for Spontaneous and Transcription-Stimulated Mitotic Recombination inSaccharomyces cerevisiae

Genetics ◽  
2002 ◽  
Vol 162 (1) ◽  
pp. 15-27 ◽  
Author(s):  
Jennifer A Freedman ◽  
Sue Jinks-Robertson
Keyword(s):  
Gene Conversion ◽  
Molecular Mechanisms ◽  
Inducible Promoter ◽  
Mitotic Recombination ◽  
Low Level ◽  
Recombination Proteins ◽  
Recombination System ◽  
Gene Conversions

AbstractThe genetic requirements for spontaneous and transcription-stimulated mitotic recombination were determined using a recombination system that employs heterochromosomal lys2 substrates that can recombine only by crossover or only by gene conversion. The substrates were fused either to a constitutive low-level promoter (pLYS) or to a highly inducible promoter (pGAL). In the case of the “conversion-only” substrates the use of heterologous promoters allowed either the donor or the recipient allele to be highly transcribed. Transcription of the donor allele stimulated gene conversions in rad50, rad51, rad54, and rad59 mutants, but not in rad52, rad55, and rad57 mutants. In contrast, transcription of the recipient allele stimulated gene conversions in rad50, rad51, rad54, rad55, rad57, and rad59 mutants, but not in rad52 mutants. Finally, transcription stimulated crossovers in rad50, rad54, and rad59 mutants, but not in rad51, rad52, rad55, and rad57 mutants. These data are considered in relation to previously proposed molecular mechanisms of transcription-stimulated recombination and in relation to the roles of the recombination proteins.

scholarly journals FREQUENCY AND DIRECTIONALITY OF GENE CONVERSION EVENTS INVOLVING THE CYC7-H3 MUTATION IN SACCHAROMYCES CEREVISIAE

Genetics ◽  
1986 ◽  
Vol 114 (2) ◽  
pp. 347-361
Author(s):  
Patricia J Pukkila ◽  
Michael D Stephens ◽  
David M Binninger ◽  
Beverly Errede
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Conversion ◽  
High Frequency ◽  
Yeast Genome ◽  
Junction Region ◽  
Sequence Comparisons ◽  
Site Specific ◽  
Site Specific Recombination ◽  
Conversion Frequency ◽  
Aberrant Segregation

ABSTRACT The CYC7-H3 mutation is a 5-kb deletion that causes overproduction of iso-2 cytochrome c. Unlike most mutations in yeast, the CYC7-H3 mutation is preferentially lost when it is involved in a gene conversion event. We have shown that cloned copies of CYC7-H3 DNA that are inserted into the yeast genome are associated with a high frequency of recombination and aberrant segregation events. Since parity in conversion frequency was observed when the extensive insertion/deletion heterozygosity at this locus was eliminated, we conclude that the CYC7-H3 sequences are inherently capable of acting as donors or recipients in gene conversion events, although they are unlikely to act as donors when they are located opposite a large heterology. DNA sequence comparisons revealed similarities between the CYC7-H3 junction region and the 2-µm circle DNA region that is involved in site-specific recombination.

scholarly journals Rearrangements occurring adjacent to a single Ty1 yeast retrotransposon in the presence and absence of full-length Ty1 transcription.

Genetics ◽  
1992 ◽  
Vol 131 (4) ◽  
pp. 833-850
Author(s):  
P R Sutton ◽  
S W Liebman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Homologous Recombination ◽  
Transposable Element ◽  
Gene Conversion ◽  
Genetic Background ◽  
Repetitive Sequences ◽  
Full Length ◽  
Yeast Genome ◽  
Yeast Saccharomyces Cerevisiae ◽  
Ty1 Retrotransposon

Abstract The structures of two unusual deletions from the yeast Saccharomyces cerevisiae are described. These deletions extend from a single Ty1 retrotransposon to an endpoint near a repetitive tRNA(Gly) gene. The deletions suggest that unique sequences flanked by two nonidentical repetitive sequences, or bordered on only one side by a transposable element, have the potential to be mobilized in the yeast genome. Models for the formation of these two unusual deletions were tested by isolating and analyzing 32 additional unusual deletions of the CYC1 region that extend from a single Ty1 retrotransposon. Unlike the most common class of deletions recovered in this region, these deletions are not attributable solely to homologous recombination among repetitive Ty1 or delta elements. They arose by two distinct mechanisms. In an SPT8 genetic background, most unusual deletions arose by transposition of a Ty1 element to a position adjacent to a tRNA(Gly) gene followed by Ty1-Ty1 recombination. In an spt8 strain, where full-length Ty1 transcription and, therefore, transposition are reduced, most deletions were due to gene conversion of a 7-kb chromosomal interval flanked by a Ty1 element and a tRNA(Gly) gene.

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