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Meiotic recombination initiation in and around retrotransposable elements in Saccharomyces cerevisiae.

Sasaki M, Tischfield SE, van Overbeek M, Keeney S - PLoS Genet. (2013)

Bottom Line: When they do, they create a risk for deleterious genome rearrangements in the germ line via recombination between non-allelic repeats.From whole-genome DSB maps and direct molecular assays, we find that DSB levels and chromatin structure within and near Tys vary widely between different elements and that local DSB suppression is not a universal feature of Ty presence.Given high strain-to-strain variability in Ty location and the high aggregate burden of Ty-proximal DSBs, we propose that meiotic recombination is an important component of host-Ty interactions and that Tys play critical roles in genome instability and evolution in both inbred and outcrossed sexual cycles.

View Article: PubMed Central - PubMed

Affiliation: Molecular Biology Program, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America.

ABSTRACT
Meiotic recombination is initiated by large numbers of developmentally programmed DNA double-strand breaks (DSBs), ranging from dozens to hundreds per cell depending on the organism. DSBs formed in single-copy sequences provoke recombination between allelic positions on homologous chromosomes, but DSBs can also form in and near repetitive elements such as retrotransposons. When they do, they create a risk for deleterious genome rearrangements in the germ line via recombination between non-allelic repeats. A prior study in budding yeast demonstrated that insertion of a Ty retrotransposon into a DSB hotspot can suppress meiotic break formation, but properties of Ty elements in their most common physiological contexts have not been addressed. Here we compile a comprehensive, high resolution map of all Ty elements in the rapidly and efficiently sporulating S. cerevisiae strain SK1 and examine DSB formation in and near these endogenous retrotransposable elements. SK1 has 30 Tys, all but one distinct from the 50 Tys in S288C, the source strain for the yeast reference genome. From whole-genome DSB maps and direct molecular assays, we find that DSB levels and chromatin structure within and near Tys vary widely between different elements and that local DSB suppression is not a universal feature of Ty presence. Surprisingly, deletion of two Ty elements weakened adjacent DSB hotspots, revealing that at least some Ty insertions promote rather than suppress nearby DSB formation. Given high strain-to-strain variability in Ty location and the high aggregate burden of Ty-proximal DSBs, we propose that meiotic recombination is an important component of host-Ty interactions and that Tys play critical roles in genome instability and evolution in both inbred and outcrossed sexual cycles.

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Deleting Ty elements increases DSB formation nearby.(A–D) Genomic DNA was isolated from meiotic cultures of a dmc1Δ strain containing the full complement of SK1 Tys and dmc1Δ strains in which either TyEST3-FAA3 or TyCGR1-SCW11 was deleted. DSBs were detected by Southern blotting and indirect end-labeling. Figures are labeled as in Figures 4C–F. Circled lower case roman numerals indicate hotspots discussed in the text. Red numerals are DSB frequencies within the bracketed regions in each of two independent cultures, corrected where appropriate for differences in transfer efficiency for the parental fragments (see Materials and Methods). Blots were stripped and rehybridized to probes from separate loci to serve as loading controls (lower panels). (A,B) DSBs around the TyEST3-FAA3 insertion site, probed from either side. (C,D) DSBs around the TyCGR1-SCW11 insertion site, probed from either side. (E) DSBs at the YCR048W hotspot (control locus) in the same samples as in panels A–D.
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pgen-1003732-g006: Deleting Ty elements increases DSB formation nearby.(A–D) Genomic DNA was isolated from meiotic cultures of a dmc1Δ strain containing the full complement of SK1 Tys and dmc1Δ strains in which either TyEST3-FAA3 or TyCGR1-SCW11 was deleted. DSBs were detected by Southern blotting and indirect end-labeling. Figures are labeled as in Figures 4C–F. Circled lower case roman numerals indicate hotspots discussed in the text. Red numerals are DSB frequencies within the bracketed regions in each of two independent cultures, corrected where appropriate for differences in transfer efficiency for the parental fragments (see Materials and Methods). Blots were stripped and rehybridized to probes from separate loci to serve as loading controls (lower panels). (A,B) DSBs around the TyEST3-FAA3 insertion site, probed from either side. (C,D) DSBs around the TyCGR1-SCW11 insertion site, probed from either side. (E) DSBs at the YCR048W hotspot (control locus) in the same samples as in panels A–D.

Mentions: To test whether natural Ty elements directly affect adjacent DSB formation, we individually deleted two Tys and compared DSB patterns with and without these elements present. As a control, we quantified DSBs in the same cultures at the YCR048W hotspot on Chr III; DSBs at this hotspot were similar between the parental and Ty deletion strains (Figure 6E).


Meiotic recombination initiation in and around retrotransposable elements in Saccharomyces cerevisiae.

Sasaki M, Tischfield SE, van Overbeek M, Keeney S - PLoS Genet. (2013)

Deleting Ty elements increases DSB formation nearby.(A–D) Genomic DNA was isolated from meiotic cultures of a dmc1Δ strain containing the full complement of SK1 Tys and dmc1Δ strains in which either TyEST3-FAA3 or TyCGR1-SCW11 was deleted. DSBs were detected by Southern blotting and indirect end-labeling. Figures are labeled as in Figures 4C–F. Circled lower case roman numerals indicate hotspots discussed in the text. Red numerals are DSB frequencies within the bracketed regions in each of two independent cultures, corrected where appropriate for differences in transfer efficiency for the parental fragments (see Materials and Methods). Blots were stripped and rehybridized to probes from separate loci to serve as loading controls (lower panels). (A,B) DSBs around the TyEST3-FAA3 insertion site, probed from either side. (C,D) DSBs around the TyCGR1-SCW11 insertion site, probed from either side. (E) DSBs at the YCR048W hotspot (control locus) in the same samples as in panels A–D.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3757047&req=5

pgen-1003732-g006: Deleting Ty elements increases DSB formation nearby.(A–D) Genomic DNA was isolated from meiotic cultures of a dmc1Δ strain containing the full complement of SK1 Tys and dmc1Δ strains in which either TyEST3-FAA3 or TyCGR1-SCW11 was deleted. DSBs were detected by Southern blotting and indirect end-labeling. Figures are labeled as in Figures 4C–F. Circled lower case roman numerals indicate hotspots discussed in the text. Red numerals are DSB frequencies within the bracketed regions in each of two independent cultures, corrected where appropriate for differences in transfer efficiency for the parental fragments (see Materials and Methods). Blots were stripped and rehybridized to probes from separate loci to serve as loading controls (lower panels). (A,B) DSBs around the TyEST3-FAA3 insertion site, probed from either side. (C,D) DSBs around the TyCGR1-SCW11 insertion site, probed from either side. (E) DSBs at the YCR048W hotspot (control locus) in the same samples as in panels A–D.
Mentions: To test whether natural Ty elements directly affect adjacent DSB formation, we individually deleted two Tys and compared DSB patterns with and without these elements present. As a control, we quantified DSBs in the same cultures at the YCR048W hotspot on Chr III; DSBs at this hotspot were similar between the parental and Ty deletion strains (Figure 6E).

Bottom Line: When they do, they create a risk for deleterious genome rearrangements in the germ line via recombination between non-allelic repeats.From whole-genome DSB maps and direct molecular assays, we find that DSB levels and chromatin structure within and near Tys vary widely between different elements and that local DSB suppression is not a universal feature of Ty presence.Given high strain-to-strain variability in Ty location and the high aggregate burden of Ty-proximal DSBs, we propose that meiotic recombination is an important component of host-Ty interactions and that Tys play critical roles in genome instability and evolution in both inbred and outcrossed sexual cycles.

View Article: PubMed Central - PubMed

Affiliation: Molecular Biology Program, Memorial Sloan-Kettering Cancer Center, New York, New York, United States of America.

ABSTRACT
Meiotic recombination is initiated by large numbers of developmentally programmed DNA double-strand breaks (DSBs), ranging from dozens to hundreds per cell depending on the organism. DSBs formed in single-copy sequences provoke recombination between allelic positions on homologous chromosomes, but DSBs can also form in and near repetitive elements such as retrotransposons. When they do, they create a risk for deleterious genome rearrangements in the germ line via recombination between non-allelic repeats. A prior study in budding yeast demonstrated that insertion of a Ty retrotransposon into a DSB hotspot can suppress meiotic break formation, but properties of Ty elements in their most common physiological contexts have not been addressed. Here we compile a comprehensive, high resolution map of all Ty elements in the rapidly and efficiently sporulating S. cerevisiae strain SK1 and examine DSB formation in and near these endogenous retrotransposable elements. SK1 has 30 Tys, all but one distinct from the 50 Tys in S288C, the source strain for the yeast reference genome. From whole-genome DSB maps and direct molecular assays, we find that DSB levels and chromatin structure within and near Tys vary widely between different elements and that local DSB suppression is not a universal feature of Ty presence. Surprisingly, deletion of two Ty elements weakened adjacent DSB hotspots, revealing that at least some Ty insertions promote rather than suppress nearby DSB formation. Given high strain-to-strain variability in Ty location and the high aggregate burden of Ty-proximal DSBs, we propose that meiotic recombination is an important component of host-Ty interactions and that Tys play critical roles in genome instability and evolution in both inbred and outcrossed sexual cycles.

Show MeSH
Related in: MedlinePlus