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| Status |
Public on Jun 09, 2021 |
| Title |
Spo11 generates gaps through concerted cuts at sites of topological stress [Top2] |
| Organisms |
Saccharomyces cerevisiae; Nakaseomyces glabratus |
| Experiment type |
Genome binding/occupancy profiling by high throughput sequencing
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| Summary |
Meiotic recombination is essential for proper meiotic chromosome segregation and fertility, and is initiated by programmed DNA double-strand breaks (DSBs) introduced by Spo11, a eukaryotic homolog of archaeal topoisomerase VIA. Here we report the discovery of hitherto uncharacterized Spo11-induced lesions, small gaps from 34 bp to several hundred bp, which are generated by coordinated pairs of DSBs (double DSBs or dDSBs). Isolation and genome-wide mapping of the resulting fragments with single base pair precision reveals enrichment at DSB hotspots but also a widely dispersed distribution covering the entire genome. We show that Spo11 prefers to cut at a sequence motif which promotes DNA bending, indicating that bendability of DNA contributes to cleavage site choice. Moreover, fragment lengths display a ~ (10.4n+3) bp periodicity, implying that Spo11 favours cleavage on the same face of underwound DNA. Consistently, dDSB signals overlap and correlate with topoisomerase II binding sites, which points to a role for topological stress and DNA crossings in break formation, and suggests a unified model for DSB and dDSB formation, in which Spo11 traps two DNA strands. Furthermore, gaps resulting from dDSBs, an estimated 20% of all initiation events, can account for full gene conversion events that are independent of both Msh2-dependent heteroduplex repair and MutLĪ³. Since non-homologous gap repair results in deletions, and ectopically re-integrated dDSB fragments result in insertions, dDSB formation represents a potential source of evolutionary diversity and pathogenic germ-line aberrations.
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| Overall design |
Chromatin immunoprecipitation (ChIP) of wild-type Saccharomyces cerevisiae (S. cer.) strains with myc-tagged (or untagged) Top2 from meiotic yeast cultures for 0 hrs - 4 hrs (t0 - t4) was performed and deep sequenced with Illumina. S. cer. cells were mixed with Candida glabrata (C. gla.) cells for calibration and downstream quantitative comparison. We are providing 4 immonoprecipitation (IP) and 4 corresponding whole cell extract (WCE) samples.
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| Contributor(s) |
Klein F, Chen D, Huang L |
| Citation(s) |
34108684 |
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| Submission date |
Mar 26, 2021 |
| Last update date |
Jun 25, 2021 |
| Contact name |
Franz Klein |
| E-mail(s) |
franz.klein@univie.ac.at
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| Organization name |
Max Perutz Labs, University of Vienna
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| Department |
Chromosome Biology
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| Street address |
Dr. Bohr-Gasse 9
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| City |
Vienna |
| ZIP/Postal code |
1030 |
| Country |
Austria |
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| Platforms (1) |
| GPL29937 |
NextSeq 550 ([Candida] glabrata; Saccharomyces cerevisiae) |
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| Samples (8)
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| This SubSeries is part of SuperSeries: |
| GSE171046 |
Spo11 generates gaps through concerted cuts at sites of topological stress |
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| Relations |
| BioProject |
PRJNA717811 |
| SRA |
SRP312382 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE169760_RAW.tar |
247.4 Mb |
(http)(custom) |
TAR (of TXT, WIG) |
| GSE169760_Top2_alignment_counts_and_calibration_factors_ASM205788v1.xlsx |
10.4 Kb |
(ftp)(http) |
XLSX |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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