Hinman AW, Yeh HY, Roelens B, Yamaya K, Woglar A, Bourbon HG, Chi P, Villeneuve AM.

Proc Natl Acad Sci U S A. 2021 Aug 17;118(33):e2109306118. doi:10.1073/pnas.2109306118

_Caenorhabditis elegans+ DSB-3 reveals conservation and divergence among protein complexes promoting meiotic double-strand breaks.

DSB-3, likely homologue of Mei4, interacts with DSB-1 and DSB-2 (likely homologues of Rec114). Together, they are interdependent for localisation to meiotic nuclei and promoting formation of crossovers. Yeast two-hybrid experiments that DSB-3 binds SPO-11 via DSB-1.

Nature. 2021 Jun;594(7864):577-582. doi:10.1038/s41586-021-03632-x

Prieler S, Chen D, Huang L, Mayrhofer E, Zsótér S, Vesely M, Mbogning J, Klein F.

Spo11 generates gaps through concerted cuts at sites of topological stress.

Spo11 has a binding preference for CN7AAGCA|TGCTTN7G and for bendable DNA. Double Spo11 cleavage generates chromosome gaps repaired by gene conversion.

“we developed ‘Protec-seq’ for the purification of end-protected DNA, which includes ChIP of Spo11, ExoV digestion, and the removal of residual 5′-tyrosyl [with hTDP2 (BPS Bioscience)] followed by deep sequencing.”

“dDSB fragments also densely cover chromosomal regions between hotspots, with approximately 31% of cuts outside of hotspots in wild-type cells and around 62% in rad50S mutants”

”The high precision of Protec-seq enabled us to identify that Spo11 has a preference for sequences that partially match a 26-nt long palindromic motif, CN7AAGCA|TGCTTN7G, centred at the cleavage axis (Fig. 2a), and a preference for C and G at position ±13 bp marking the border of the footprint of Spo1133.”

“Dividing the preferred fragment lengths into n helical turns leads to helix lengths above 10.4 bp, indicative of underwound DNA. [...] DNA at promoters is underwound or negatively supercoiled [...] In both wild-type and rad50S cells, (d)DSB levels correlate positively with the corresponding transcriptional stress.”

”As an evolutionary relative of a type IIB topoisomerase, the Spo11 complex may require DNA crossings for the cleavage reaction, which are likely to form at promoters known to accumulate negative supercoils. [...] topoisomerase II (Top2) prefers to bind to DNA crossings under superhelical stress [...] robust [Top2] peaks accumulate at nearly all dDSB sites by the time of DSB formation [...] the occupancy of Top2 increases moderately with higher transcriptional stress at (divergent and tandem) promoters, but not at convergent sites”

“we tested whether gaps that result from dDSBs could account for the observed 6:2 [gene conversion] events by determining the set of dDSB fragments that fully overlap a 6:2 event as an indicator for the local probability of gap formation. The 6:2 event distribution is significantly shifted to genomic positions with higher dDSB gap probability, which indicates that dDSB gaps are enriched at full gene conversion sites and of sufficient length in both wild-type strains and rad50S mutants, as opposed to spo11Y135F mutants”

Johnson D, Crawford M, Cooper T, Claeys Bouuaert C, Keeney S, Llorente B, Garcia V, Neale MJ.

Concerted cutting by Spo11 illuminates meiotic DNA break mechanics.

Nature. 2021 Jun;594(7864):572-576. doi:10.1038/s41586-021-03389-3

Pairs of double strand breaks created by Spo11 causes loss of short oligonucleotides, creating a cap repaired by homologous recombination.

“High-resolution analysis of deproteinized Spo11-oligos showed [a ladder of] periodicity of around 10 nt [that] ranges in length from around 33 nt to more than 100 nt” “The ladder also arose in rad50S-mutant and Mre11-nuclease-defective strains of S. cerevisiae, which, like the sae2∆ mutant, cannot remove Spo11 from DSB ends. Notably, the ladder depended on the catalytic activity of Spo11” “we refer to this ladder as hyperlocalized ‘Spo11 double cuts’ (Spo11-DCs).”

“because Spo11-DCs of less than 30 nt in length are not detected on gels and are depleted from filtered Spo11-oligo libraries, we propose that adjacent Spo11 complexes that are capable of making double cuts must, in vivo, interact with DNA from the same direction, thereby generating a ladder of Spo11-DCs with periodicity dictated by the helical pitch of DNA (around 10.5 bp).”

“the global frequency of Spo11-DCs was disproportionately associated with regions of stronger Spo11 activity”

“Deep sequencing of meiotic progeny identifies recombination scars that are consistent with repair initiated from gaps generated by adjacent Spo11 DSBs.”

“We envision a mechanism in which multiple Spo11 proteins assemble with other pro-DSB factors to create a platform that enables concerted Spo11-DSB formation”

Bell AD, Mello CJ, Nemesh J, Brumbaugh SA, Wysoker A, McCarroll SA.

Nature. 2020 Jul;583(7815):259-264. doi:10.1038/s41586-020-2347-0

Insights into variation in meiosis from 31,228 human sperm genomes.

“We identified 813,122 crossovers in the 31,228 gamete genomes.” “Gametes with fewer crossovers in half of their genome tended to have fewer crossovers in the other half of their genome.” “Large concentrations of crossovers in distal regions.” “The sex chromosomes and acrocentric chromosomes had the highest rates of aneuploidy.”

HELLS and PRDM9 form a pioneer complex to open chromatin at meiotic recombination hot spots.

Spruce C, Dlamini S, Ananda G, Bronkema N, Tian H, Paigen K, Carter GW, Baker CL.

Genes Dev. 2020 Mar 1;34(5-6):398-412. doi:10.1101/gad.333542.119

“[One] chromatin states [...] was enriched for PRDM9-binding sites.” (H3K4me3 > H3K4me1 > H3K36me3 > H3K9ac) “The majority [of the] locations overlap with previously reported locations of B6 meiosis-specific DSBs and PRDM9-dependent H3K4me3 modification.” “Interestingly, compared with other phyla, we did not detect H2A.Z at hot spots in mice.” “At PRDM9-dependent H3K4me3 sites, we detected increased DNA accessibility [ATAC-seq] overlapping with PRDM9 motif locations.” Changing the DNA binding specificity of PRDM9 moved the whole epigenetic mark to new locations. Testis-conditional KO of Hells relocates meiotic DSBs (identified by DMC1 binding) to promoter regions (determined as such by annotation and by chromatin state). The hotspot chromatin state is reduced in Hells mutants, although PRDM9 is still present. “These data show that HELLS is required for establishment of the epigenomic state and chromatin accessibility at hot spots.” Hells interacts with a “PRDM9 variant that lacks the DNA-binding domain (PRDM9ΔZF) [...], suggesting that this interaction is independent of PRDM9-directed DNA binding.” “Critically, upon loss of HELLS, PRDM9C binding at hot spots was reduced to background levels (Fig. 6H). Together, these data show that PRDM9 and HELLS form a complex in vivo and, by extension, suggests that active chromatin remodeling is required for robust PRDM9 binding at hot spots.”

Singhal S, Leffler EM, Sannareddy K, Turner I, Venn O, Hooper DM, Strand AI, Li Q, Raney B, Balakrishnan CN, Griffith SC, McVean G, Przeworski M.

Science. 2015 Nov 20;350(6263):928-32. doi:10.1126/science.aad0843

Stable recombination hotspots in birds.

Recombination maps obtained from genome sequences of 24 zebra finches and 20 long-tail finches, in which SNP Haplotypes were inferred from phase-informative reads and family phasing. Recombination rates estimated as ρ = 26.2/kb and 14.0/kb, respectively (0.14 cM/Mb in both species). 3—4000 Hotspots “operationally defined them as regions that are at least 2 kb in length; have at least five times the background recombination rate as estimated across the 80 kb of sequence surrounding the region; and are statistically supported as hotspots by a likelihood ratio test” (18). “73% of zebra finch hotspots (...) were detected as shared between the two species.” “Hotspots in the zebra finch and long-tailed finch genomes are enriched near transcription start sites (TSSs), transcription stop sites (TESs), and CpG islands (CGIs), with close to half of all hotspots occurring within 3 kb of one of these features.” “Median recombination rates across and within chromosomes vary over nearly six orders of magnitude (...) with regions of elevated recombination near telomeres and large intervening deserts.

Shibuya H, Ishiguro K, Watanabe Y.

Nat Cell Biol. 2014 Feb;16(2):145-56. doi:10.1038/ncb2896

The TRF1-binding protein TERB1 promotes chromosome movement and telomere rigidity in meiosis.

Stabilises mechanical transduction by the SUN-KASH complex to the meiotic chromosomes.