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Genome-wide high-resolution mapping of UV-induced mitotic recombination events in Saccharomyces cerevisiae.

Yin Y, Petes TD - PLoS Genet. (2013)

Bottom Line: Mitotic recombination between homologous chromosomes can result in loss of heterozygosity (LOH).UV doses that have little effect on the viability of diploid cells stimulate crossovers more than 1000-fold in wild-type cells.Genome-wide mapping of about 380 unselected crossovers, break-induced replication (BIR) events, and gene conversions shows that UV-induced recombination events occur throughout the genome without pronounced hotspots, although the ribosomal RNA gene cluster has a significantly lower frequency of crossovers.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Microbiology and University Program in Genetics and Genomics, Duke University Medical Center, Durham, North Carolina, United States of America.

ABSTRACT
In the yeast Saccharomyces cerevisiae and most other eukaryotes, mitotic recombination is important for the repair of double-stranded DNA breaks (DSBs). Mitotic recombination between homologous chromosomes can result in loss of heterozygosity (LOH). In this study, LOH events induced by ultraviolet (UV) light are mapped throughout the genome to a resolution of about 1 kb using single-nucleotide polymorphism (SNP) microarrays. UV doses that have little effect on the viability of diploid cells stimulate crossovers more than 1000-fold in wild-type cells. In addition, UV stimulates recombination in G1-synchronized cells about 10-fold more efficiently than in G2-synchronized cells. Importantly, at high doses of UV, most conversion events reflect the repair of two sister chromatids that are broken at approximately the same position whereas at low doses, most conversion events reflect the repair of a single broken chromatid. Genome-wide mapping of about 380 unselected crossovers, break-induced replication (BIR) events, and gene conversions shows that UV-induced recombination events occur throughout the genome without pronounced hotspots, although the ribosomal RNA gene cluster has a significantly lower frequency of crossovers.

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Related in: MedlinePlus

Mapping crossovers on chromosome V by SNP arrays.Genomic DNA was isolated from the red and white sectors of a sectored colony derived from UV-treated cells. The ratio of hybridization of SNP-specific oligonucleotides (relative to the hybridization levels of DNA from a fully heterozygous strain) for each sample was measured (details in text) and is shown on the Y axis. A hybridization level of about 1 indicates that the strain was heterozygous. The X axis shows the SGD coordinates of chromosome V beginning at the left telomere. Black and red lines indicate the normalized hybridization ratio to the YJM789- and W303a-specific oligonucleotides, respectively. The crossover and conversions associated with the sectors are diagrammed with the upper and lower panels showing patterns of LOH in the red and white sectors, respectively. A. Low-resolution depiction of the LOH events in the red and white sectors. The transition is at about SGD coordinate 105 kb. B. High-resolution depiction of LOH events (same event as shown in Figure 4A). Black squares show hybridization to YJM789-specific oligonucleotides and red diamonds show hybridization to W303a-specific oligonucleotides. The red sector has a single transition between heterozygous and homozygous SNPs, whereas the white sector has three transitions. The pattern of transitions is consistent with a 3∶1/4∶0 hybrid tract associated with a crossover. C. Summary of patterns of heterozygous and homozygous markers with the top line showing the red sector and the bottom line showing the white sector. The black, red, and green lines indicate regions homozygous for YJM789-derived SNPs, homozygous for W303a-derived SNPs, and heterozygous regions, respectively. The orange circles show the position of the centromeres. The lengths of the line segments showing the LOH region associated with the crossover and the heterozygous region centromere-proximal to the crossover are not shown to scale.
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pgen-1003894-g004: Mapping crossovers on chromosome V by SNP arrays.Genomic DNA was isolated from the red and white sectors of a sectored colony derived from UV-treated cells. The ratio of hybridization of SNP-specific oligonucleotides (relative to the hybridization levels of DNA from a fully heterozygous strain) for each sample was measured (details in text) and is shown on the Y axis. A hybridization level of about 1 indicates that the strain was heterozygous. The X axis shows the SGD coordinates of chromosome V beginning at the left telomere. Black and red lines indicate the normalized hybridization ratio to the YJM789- and W303a-specific oligonucleotides, respectively. The crossover and conversions associated with the sectors are diagrammed with the upper and lower panels showing patterns of LOH in the red and white sectors, respectively. A. Low-resolution depiction of the LOH events in the red and white sectors. The transition is at about SGD coordinate 105 kb. B. High-resolution depiction of LOH events (same event as shown in Figure 4A). Black squares show hybridization to YJM789-specific oligonucleotides and red diamonds show hybridization to W303a-specific oligonucleotides. The red sector has a single transition between heterozygous and homozygous SNPs, whereas the white sector has three transitions. The pattern of transitions is consistent with a 3∶1/4∶0 hybrid tract associated with a crossover. C. Summary of patterns of heterozygous and homozygous markers with the top line showing the red sector and the bottom line showing the white sector. The black, red, and green lines indicate regions homozygous for YJM789-derived SNPs, homozygous for W303a-derived SNPs, and heterozygous regions, respectively. The orange circles show the position of the centromeres. The lengths of the line segments showing the LOH region associated with the crossover and the heterozygous region centromere-proximal to the crossover are not shown to scale.

Mentions: Figure 4 shows the analysis of one red/white sectored colony (59RW). In this figure, we show the normalized ratio of hybridization of genomic sequences to W303a- and YJM789-specific oligonucleotides on chromosome V with red lines and black lines, respectively; CEN5 is located near coordinate 152 kb. In the top part of Figure 4A, we depict the pattern of hybridization of genomic DNA isolated from the red sector. The ratio of hybridization is about 1 for all SNPs from coordinate 105 kb to the right telomere, indicating that SNPs in this region are heterozygous. In the red sector, SNPs centromere-distal to coordinate 105 kb on the left arm are homozygous for the W303a-derived SNPs whereas the genomic DNA from the white sector becomes homozygous at approximately the same position for YJM789-derived SNPs. In Figure 4B, the same recombination event is depicted at higher resolution; each square and diamond shows the level of hybridization to an individual YJM789-specific or a W303a-specific SNP, respectively. As shown in this figure, the red sector has a single transition between heterozygous and homozygous SNPs whereas the white sector has three transitions. The pattern of these transitions indicates that the crossover is associated with a 3∶1/4∶0 hybrid conversion tract.


Genome-wide high-resolution mapping of UV-induced mitotic recombination events in Saccharomyces cerevisiae.

Yin Y, Petes TD - PLoS Genet. (2013)

Mapping crossovers on chromosome V by SNP arrays.Genomic DNA was isolated from the red and white sectors of a sectored colony derived from UV-treated cells. The ratio of hybridization of SNP-specific oligonucleotides (relative to the hybridization levels of DNA from a fully heterozygous strain) for each sample was measured (details in text) and is shown on the Y axis. A hybridization level of about 1 indicates that the strain was heterozygous. The X axis shows the SGD coordinates of chromosome V beginning at the left telomere. Black and red lines indicate the normalized hybridization ratio to the YJM789- and W303a-specific oligonucleotides, respectively. The crossover and conversions associated with the sectors are diagrammed with the upper and lower panels showing patterns of LOH in the red and white sectors, respectively. A. Low-resolution depiction of the LOH events in the red and white sectors. The transition is at about SGD coordinate 105 kb. B. High-resolution depiction of LOH events (same event as shown in Figure 4A). Black squares show hybridization to YJM789-specific oligonucleotides and red diamonds show hybridization to W303a-specific oligonucleotides. The red sector has a single transition between heterozygous and homozygous SNPs, whereas the white sector has three transitions. The pattern of transitions is consistent with a 3∶1/4∶0 hybrid tract associated with a crossover. C. Summary of patterns of heterozygous and homozygous markers with the top line showing the red sector and the bottom line showing the white sector. The black, red, and green lines indicate regions homozygous for YJM789-derived SNPs, homozygous for W303a-derived SNPs, and heterozygous regions, respectively. The orange circles show the position of the centromeres. The lengths of the line segments showing the LOH region associated with the crossover and the heterozygous region centromere-proximal to the crossover are not shown to scale.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3814309&req=5

pgen-1003894-g004: Mapping crossovers on chromosome V by SNP arrays.Genomic DNA was isolated from the red and white sectors of a sectored colony derived from UV-treated cells. The ratio of hybridization of SNP-specific oligonucleotides (relative to the hybridization levels of DNA from a fully heterozygous strain) for each sample was measured (details in text) and is shown on the Y axis. A hybridization level of about 1 indicates that the strain was heterozygous. The X axis shows the SGD coordinates of chromosome V beginning at the left telomere. Black and red lines indicate the normalized hybridization ratio to the YJM789- and W303a-specific oligonucleotides, respectively. The crossover and conversions associated with the sectors are diagrammed with the upper and lower panels showing patterns of LOH in the red and white sectors, respectively. A. Low-resolution depiction of the LOH events in the red and white sectors. The transition is at about SGD coordinate 105 kb. B. High-resolution depiction of LOH events (same event as shown in Figure 4A). Black squares show hybridization to YJM789-specific oligonucleotides and red diamonds show hybridization to W303a-specific oligonucleotides. The red sector has a single transition between heterozygous and homozygous SNPs, whereas the white sector has three transitions. The pattern of transitions is consistent with a 3∶1/4∶0 hybrid tract associated with a crossover. C. Summary of patterns of heterozygous and homozygous markers with the top line showing the red sector and the bottom line showing the white sector. The black, red, and green lines indicate regions homozygous for YJM789-derived SNPs, homozygous for W303a-derived SNPs, and heterozygous regions, respectively. The orange circles show the position of the centromeres. The lengths of the line segments showing the LOH region associated with the crossover and the heterozygous region centromere-proximal to the crossover are not shown to scale.
Mentions: Figure 4 shows the analysis of one red/white sectored colony (59RW). In this figure, we show the normalized ratio of hybridization of genomic sequences to W303a- and YJM789-specific oligonucleotides on chromosome V with red lines and black lines, respectively; CEN5 is located near coordinate 152 kb. In the top part of Figure 4A, we depict the pattern of hybridization of genomic DNA isolated from the red sector. The ratio of hybridization is about 1 for all SNPs from coordinate 105 kb to the right telomere, indicating that SNPs in this region are heterozygous. In the red sector, SNPs centromere-distal to coordinate 105 kb on the left arm are homozygous for the W303a-derived SNPs whereas the genomic DNA from the white sector becomes homozygous at approximately the same position for YJM789-derived SNPs. In Figure 4B, the same recombination event is depicted at higher resolution; each square and diamond shows the level of hybridization to an individual YJM789-specific or a W303a-specific SNP, respectively. As shown in this figure, the red sector has a single transition between heterozygous and homozygous SNPs whereas the white sector has three transitions. The pattern of these transitions indicates that the crossover is associated with a 3∶1/4∶0 hybrid conversion tract.

Bottom Line: Mitotic recombination between homologous chromosomes can result in loss of heterozygosity (LOH).UV doses that have little effect on the viability of diploid cells stimulate crossovers more than 1000-fold in wild-type cells.Genome-wide mapping of about 380 unselected crossovers, break-induced replication (BIR) events, and gene conversions shows that UV-induced recombination events occur throughout the genome without pronounced hotspots, although the ribosomal RNA gene cluster has a significantly lower frequency of crossovers.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Microbiology and University Program in Genetics and Genomics, Duke University Medical Center, Durham, North Carolina, United States of America.

ABSTRACT
In the yeast Saccharomyces cerevisiae and most other eukaryotes, mitotic recombination is important for the repair of double-stranded DNA breaks (DSBs). Mitotic recombination between homologous chromosomes can result in loss of heterozygosity (LOH). In this study, LOH events induced by ultraviolet (UV) light are mapped throughout the genome to a resolution of about 1 kb using single-nucleotide polymorphism (SNP) microarrays. UV doses that have little effect on the viability of diploid cells stimulate crossovers more than 1000-fold in wild-type cells. In addition, UV stimulates recombination in G1-synchronized cells about 10-fold more efficiently than in G2-synchronized cells. Importantly, at high doses of UV, most conversion events reflect the repair of two sister chromatids that are broken at approximately the same position whereas at low doses, most conversion events reflect the repair of a single broken chromatid. Genome-wide mapping of about 380 unselected crossovers, break-induced replication (BIR) events, and gene conversions shows that UV-induced recombination events occur throughout the genome without pronounced hotspots, although the ribosomal RNA gene cluster has a significantly lower frequency of crossovers.

Show MeSH
Related in: MedlinePlus