Exo1 protects DNA nicks from ligation to promote crossover formation during meiosis.

In most sexually reproducing organisms crossing over between chromosome homologs during meiosis is essential to produce haploid gametes. Most crossovers that form in meiosis in budding yeast result from the biased resolution of double Holliday junction (dHJ) intermediates. This dHJ resolution step involves the actions of Rad2/XPG family nuclease Exo1 and the Mlh1-Mlh3 mismatch repair endonuclease. Here, we provide genetic evidence in baker's yeast that Exo1 promotes meiotic crossing over by protecting DNA nicks from ligation. We found that structural elements in Exo1 that interact with DNA, such as those required for the bending of DNA during nick/flap recognition, are critical for its role in crossing over. Consistent with these observations, meiotic expression of the Rad2/XPG family member Rad27 partially rescued the crossover defect in exo1 null mutants, and meiotic overexpression of Cdc9 ligase reduced the crossover levels of exo1 DNA-binding mutants to levels that approached the exo1 null. In addition, our work identified a role for Exo1 in crossover interference. Together, these studies provide experimental evidence for Exo1-protected nicks being critical for the formation of meiotic crossovers and their distribution.

Human AAA+ ATPase FIGNL1 suppresses RAD51-mediated ultra-fine bridge formation.

RAD51 filament is crucial for the homology-dependent repair of DNA double-strand breaks and stalled DNA replication fork protection. Positive and negative regulators control RAD51 filament assembly and disassembly. RAD51 is vital for genome integrity but excessive accumulation of RAD51 on chromatin causes genome instability and growth defects. However, the detailed mechanism underlying RAD51 disassembly by negative regulators and the physiological consequence of abnormal RAD51 persistence remain largely unknown. Here, we report the role of the human AAA+ ATPase FIGNL1 in suppressing a novel type of RAD51-mediated genome instability. FIGNL1 knockout human cells were defective in RAD51 dissociation after replication fork restart and accumulated ultra-fine chromosome bridges (UFBs), whose formation depends on RAD51 rather than replication fork stalling. FIGNL1 suppresses homologous recombination intermediate-like UFBs generated between sister chromatids at genomic loci with repeated sequences such as telomeres and centromeres. These data suggest that RAD51 persistence per se induces the formation of unresolved linkage between sister chromatids resulting in catastrophic genome instability. FIGNL1 facilitates post-replicative disassembly of RAD51 filament to suppress abnormal recombination intermediates and UFBs. These findings implicate FIGNL1 as a key factor required for active RAD51 removal after processing of stalled replication forks, which is essential to maintain genome stability.

RPA interacts with Rad52 to promote meiotic crossover and noncrossover recombination.

Meiotic recombination is initiated by programmed double-strand breaks (DSBs). Studies in Saccharomyces cerevisiae have shown that, following rapid resection to generate 3' single-stranded DNA (ssDNA) tails, one DSB end engages a homolog partner chromatid and is extended by DNA synthesis, whereas the other end remains associated with its sister. Then, after regulated differentiation into crossover- and noncrossover-fated types, the second DSB end participates in the reaction by strand annealing with the extended first end, along both pathways. This second-end capture is dependent on Rad52, presumably via its known capacity to anneal two ssDNAs. Here, using physical analysis of DNA recombination, we demonstrate that this process is dependent on direct interaction of Rad52 with the ssDNA binding protein, replication protein A (RPA). Furthermore, the absence of this Rad52-RPA joint activity results in a cytologically-prominent RPA spike, which emerges from the homolog axes at sites of crossovers during the pachytene stage of the meiotic prophase. Our findings suggest that this spike represents the DSB end of a broken chromatid caused by either the displaced leading DSB end or the second DSB end, which has been unable to engage with the partner homolog-associated ssDNA. These and other results imply a close correspondence between Rad52-RPA roles in meiotic recombination and mitotic DSB repair.

Mutational analysis of Mei5, a subunit of Mei5-Sae3 complex, in Dmc1-mediated recombination during yeast meiosis.

Interhomolog recombination in meiosis is mediated by the Dmc1 recombinase. The Mei5-Sae3 complex of Saccharomyces cerevisiae promotes Dmc1 assembly and functions with Dmc1 for homology-mediated repair of meiotic DNA double-strand breaks. How Mei5-Sae3 facilitates Dmc1 assembly remains poorly understood. In this study, we created and characterized several mei5 mutants featuring the amino acid substitutions of basic residues. We found that Arg97 of Mei5, conserved in its ortholog, SFR1 (complex with SWI5), RAD51 mediator, in humans and other organisms, is critical for complex formation with Sae3 for Dmc1 assembly. Moreover, the substitution of either Arg117 or Lys133 with Ala in Mei5 resulted in the production of a C-terminal truncated Mei5 protein during yeast meiosis. Notably, the shorter Mei5-R117A protein was observed in meiotic cells but not in mitotic cells when expressed, suggesting a unique regulation of Dmc1-mediated recombination by posttranslational processing of Mei5-Sae3.

DNA double-strand breaks regulate the cleavage-independent release of Rec8-cohesin during yeast meiosis.

The mitotic cohesin complex necessary for sister chromatid cohesion and chromatin loop formation shows local and global association to chromosomes in response to DNA double-strand breaks (DSBs). Here, by genome-wide binding analysis of the meiotic cohesin with Rec8, we found that the Rec8-localization profile along chromosomes is altered from middle to late meiotic prophase I with cleavage-independent dissociation. Each Rec8-binding site on the chromosome axis follows a unique alternation pattern with dissociation and probably association. Centromeres showed altered Rec8 binding in late prophase I relative to mid-prophase I, implying chromosome remodeling of the regions. Rec8 dissociation ratio per chromosome is correlated well with meiotic DSB density. Indeed, the spo11 mutant deficient in meiotic DSB formation did not change the distribution of Rec8 along chromosomes in late meiotic prophase I. These suggest the presence of a meiosis-specific regulatory pathway for the global binding of Rec8-cohesin in response to DSBs.

Fanconi anemia-associated mutation in RAD51 compromises the coordinated action of DNA-binding and ATPase activities.

Fanconi anemia (FA) is a rare genetic disease caused by a defect in DNA repair pathway for DNA interstrand crosslinks. These crosslinks can potentially impede the progression of the DNA replication fork, consequently leading to DNA double-strand breaks. Heterozygous RAD51-Q242R mutation has been reported to cause FA-like symptoms. However, the molecular defect of RAD51 underlying the disease is largely unknown. In this study, we conducted a biochemical analysis of RAD51-Q242R protein, revealing notable deficiencies in its DNA-dependent ATPase activity and its ATP-dependent regulation of DNA-binding activity. Interestingly, although RAD51-Q242R exhibited the filament instability and lacked the ability to form displacement loop, it efficiently stimulated the formation of displacement loops mediated by wild-type RAD51. These findings facilitate understanding of the biochemical properties of the mutant protein and how RAD51 works in the FA patient cells.

The synaptonemal complex central region modulates crossover pathways and feedback control of meiotic double-strand break formation.

The synaptonemal complex (SC) is a proteinaceous structure that mediates homolog engagement and genetic recombination during meiosis. In budding yeast, Zip-Mer-Msh (ZMM) proteins promote crossover (CO) formation and initiate SC formation. During SC elongation, the SUMOylated SC component Ecm11 and the Ecm11-interacting protein Gmc2 facilitate the polymerization of Zip1, an SC central region component. Through physical recombination, cytological, and genetic analyses, we found that ecm11 and gmc2 mutants exhibit chromosome-specific defects in meiotic recombination. CO frequencies on a short chromosome (chromosome III) were reduced, whereas CO and non-crossover frequencies on a long chromosome (chromosome VII) were elevated. Further, in ecm11 and gmc2 mutants, more double-strand breaks (DSBs) were formed on a long chromosome during late prophase I, implying that the Ecm11-Gmc2 (EG) complex is involved in the homeostatic regulation of DSB formation. The EG complex may participate in joint molecule (JM) processing and/or double-Holliday junction resolution for ZMM-dependent CO-designated recombination. Absence of the EG complex ameliorated the JM-processing defect in zmm mutants, suggesting a role for the EG complex in suppressing ZMM-independent recombination. Our results suggest that the SC central region functions as a compartment for sequestering recombination-associated proteins to regulate meiosis specificity during recombination.

Clinical study of preoperative skeletal muscle mass as a predictor of physical performance recovery following palliative surgery for spinal metastases.

BACKGROUND: Surgical treatment of spinal metastases has been associated with high morbidity and mortality in patients with sarcopenia based on low skeletal muscle mass. We assessed physical performance using the Eastern Cooperative Oncology Group performance status scale and the Barthel Index on the 30th day after palliative surgery for spinal metastases and investigated the effectiveness of surgery according to sarcopenia assessed by skeletal muscle mass. METHODS: We retrospectively analyzed 78 consecutive patients with thoracic and lumbar spinal metastases who underwent palliative surgery. The value of the area of the psoas major muscle at the L3 level normalized by the vertebral area was divided into first, middle, and third tertiles. Clinical variables were compared by tertile. Variables affecting the 30-day good performance status were investigated with univariate and multivariate analyses. RESULTS: The 30-day morbidity rates were 50%, 38.5%, and 15.4% by tertile. The 30-day mortality rate was 2%; all were in the first tertile. Good preoperative performance status scores were seen in 15.4% of first and 50% of third tertile patients. Postoperatively, the performance status improved in all groups, with 30.8%, 65.4%, and 92.3% by tertile. Multivariate regression analysis revealed that a good preoperative performance status (OR: 15.50, 95% CI: 1.610-149.00, P < 0.05) and the value of the area of the psoas major muscle at the L3 level normalized by the vertebral area not in the first tertile (OR: 0.22, 95% CI: 0.06-0.82, P < 0.05) were significant predictors of a good postoperative 30-day performance status. CONCLUSIONS: A good preoperative performance status and exclusion from the first tertile were clinical factors predicting a good postoperative 30-day performance status. In patients with large psoas muscle mass (third tertile), a good 30-day performance status can be expected after surgery, suggesting that surgery in this population should be pursued aggressively.

Singlet Oxygen Generation Driven by Sulfide Ligand Exchange on Porphyrin-Gold Nanoparticle Conjugates.

Here, we report a switching method of singlet oxygen (1O2) generation based on the adsorption/desorption of porphyrins to gold nanoparticles driven by sulfide (thiol or disulfide) compounds. The generation of 1O2 by photosensitization is effectively suppressed by the gold nanoparticles and can be restored by a sulfide ligand exchange reaction. The on/off ratio of 1O2 quantum yield (ΦΔ) reached 7.4. By examining various incoming sulfide compounds, it was found that the ligand exchange reaction on the gold nanoparticle surface could be thermodynamically or kinetically controlled. The remaining gold nanoparticles in the system still suppress the generation of 1O2, which can be precipitated out simultaneously with porphyrin desorption by the proper polarity choice of the incoming sulfide to restore the 1O2 generation.

Dual roles of yeast Rad51 N-terminal domain in repairing DNA double-strand breaks.

Highly toxic DNA double-strand breaks (DSBs) readily trigger the DNA damage response (DDR) in cells, which delays cell cycle progression to ensure proper DSB repair. In Saccharomyces cerevisiae, mitotic S phase (20-30 min) is lengthened upon DNA damage. During meiosis, Spo11-induced DSB onset and repair lasts up to 5 h. We report that the NH2-terminal domain (NTD; residues 1-66) of Rad51 has dual functions for repairing DSBs during vegetative growth and meiosis. Firstly, Rad51-NTD exhibits autonomous expression-enhancing activity for high-level production of native Rad51 and when fused to exogenous ß-galactosidase in vivo. Secondly, Rad51-NTD is an S/T-Q cluster domain (SCD) harboring three putative Mec1/Tel1 target sites. Mec1/Tel1-dependent phosphorylation antagonizes the proteasomal degradation pathway, increasing the half-life of Rad51 from ∼30 min to ≥180 min. Our results evidence a direct link between homologous recombination and DDR modulated by Rad51 homeostasis.

Tension Pneumocephalus Associated with Negative Pressure Wound Therapy with Instillation and Dwell Time for Methicillin-resistant Staphylococcus aureus Infection After Spinal Deformity Surgery.

PURPOSE: Surgical site infection (SSI) is a serious complication after spine surgery. Recently, it has become possible to perform negative pressure wound therapy with instillation and dwell time (NPWTi-d) for postoperative infected wounds. We report the first rare case of symptomatic pneumoencephalopathy following NPWTi-d for methicillin-resistant Staphylococcus aureus (MRSA) infection after spinal deformity surgery. METHODS: Retrospective review of a patient's medical record and imaging. RESULTS: A 77-year-old female patient underwent posterior corrective fixation with no intraoperative complications. On the 10th postoperative day, SSI was diagnosed, and debridement was performed. Since MRSA was detected in the wound culture, and a prolonged inflammatory reaction was observed, NPWTi-d was started to preserve the instrumentation. Gradually, good granulation was observed, and the extensive soft tissue defect decreased. On the 29th day after the start of NPWTi-d, the patient experienced sudden headache and neck pain while standing, and head computed tomography led to the diagnosis of symptomatic pneumoencephalopathy. NPWTi-d was discontinued, and when surgery was performed to close the wound, dural injury was found, which was not present at the time of the initial surgery, and dural repair was performed. After 2 weeks of bed rest, the patient's pneumoencephalopathy improved. Three years have passed since the surgery, and no recurrence of cerebrospinal fluid leakage or infection has been observed. CONCLUSIONS: Although NPWTi-d is a useful treatment for SSI, it is always necessary to pay attention to the development of pneumoencephalopathy and promptly diagnose and treat it because of the risk of life-threatening complications.

Meiosis-specific prophase-like pathway controls cleavage-independent release of cohesin by Wapl phosphorylation.

Sister chromatid cohesion on chromosome arms is essential for the segregation of homologous chromosomes during meiosis I while it is dispensable for sister chromatid separation during mitosis. It was assumed that, unlike the situation in mitosis, chromosome arms retain cohesion prior to onset of anaphase-I. Paradoxically, reduced immunostaining signals of meiosis-specific cohesin, including the kleisin Rec8, were observed on chromosomes during late prophase-I of budding yeast. This decrease is seen in the absence of Rec8 cleavage and depends on condensin-mediated recruitment of Polo-like kinase (PLK/Cdc5). In this study, we confirmed that this release indeed accompanies the dissociation of acetylated Smc3 as well as Rec8 from meiotic chromosomes during late prophase-I. This release requires, in addition to PLK, the cohesin regulator, Wapl (Rad61/Wpl1 in yeast), and Dbf4-dependent Cdc7 kinase (DDK). Meiosis-specific phosphorylation of Rad61/Wpl1 and Rec8 by PLK and DDK collaboratively promote this release. This process is similar to the vertebrate "prophase" pathway for cohesin release during G2 phase and pro-metaphase. In yeast, meiotic cohesin release coincides with PLK-dependent compaction of chromosomes in late meiotic prophase-I. We suggest that yeast uses this highly regulated cleavage-independent pathway to remove cohesin during late prophase-I to facilitate morphogenesis of condensed metaphase-I chromosomes.

Minimally Invasive Spinal Treatment (MIST)-A New Concept in the Treatment of Spinal Diseases: A Narrative Review.

In the past two decades, minimally invasive spine surgery (MISS) techniques have been developed for spinal surgery. Historically, minimizing invasiveness in decompression surgery was initially reported as a MISS technique. In recent years, MISS techniques have also been applied for spinal stabilization techniques, which were defined as minimally invasive spine stabilization (MISt), including percutaneous pedicle screws (PPS) fixation, lateral lumbar interbody fusion, balloon kyphoplasty, percutaneous vertebroplasty, cortical bone trajectory, and cervical total disc replacement. These MISS techniques typically provide many advantages such as preservation of paraspinal musculature, less blood loss, a shorter operative time, less postoperative pain, and a lower infection rate as well as being more cost-effective compared to traditional open techniques. However, even MISS techniques are associated with several limitations including technical difficulty, training opportunities, surgical cost, equipment cost, and radiation exposure. These downsides of surgical treatments make conservative treatments more feasible option. In the future, medicine must become "minimally invasive" in the broadest sense-for all patients, conventional surgeries, medical personnel, hospital management, nursing care, and the medical economy. As a new framework for the treatment of spinal diseases, the concept of minimally invasive spinal treatment (MIST) has been proposed.

Structural dynamics of the chromo-shadow domain and chromodomain of HP1 bound to histone H3K9 methylated peptide, as measured by site-directed spin-labeling EPR spectroscopy.

The structural dynamics of the chromo-shadow domain (CSD) and chromodomain (CD) of human HP1 proteins essential for heterochromatin formation were investigated at the nanosecond and nanometer scales by site-directed spin labeling electron paramagnetic resonance and pulsed double resonance spectroscopy. Distance measurements showed that the spin-labeled CSD of human HP1α and HP1γ tightly dimerizes. Unlike CD-CD interaction observed in fission yeast HP1 in an inactivated state (Canzio et al., 2013), the two CDs of HP1α and HP1γ were spatially separated from each other, dynamically mobile, and ready for a Brownian search for H3K9-tri-methyl(me3) on histones. Complex formation of the CD with H3K9me3 slowed dynamics of the domain due to a decreased diffusion constant. CSD mobility was significantly (∼1.3-fold) lower in full-length HP1α than in HP1γ, suggesting that the immobilized conformation of human HP1α shows an auto-inactivated state. Differential properties of HP1α and HP1γ to form the inactive conformation could be relevant to its physiological role in the heterochromatin formation in a cell.

Enhanced processivity of Dnmt1 by monoubiquitinated histone H3.

DNA methylation controls gene expression, and once established, DNA methylation patterns are faithfully copied during DNA replication by the maintenance DNA methyltransferase Dnmt1. In vivo, Dnmt1 interacts with Uhrf1, which recognizes hemimethylated CpGs. Recently, we reported that Uhrf1-catalyzed K18- and K23-ubiquitinated histone H3 binds to the N-terminal region (the replication focus targeting sequence, RFTS) of Dnmt1 to stimulate its methyltransferase activity. However, it is not yet fully understood how ubiquitinated histone H3 stimulates Dnmt1 activity. Here, we show that monoubiquitinated histone H3 stimulates Dnmt1 activity toward DNA with multiple hemimethylated CpGs but not toward DNA with only a single hemimethylated CpG, suggesting an influence of ubiquitination on the processivity of Dnmt1. The Dnmt1 activity stimulated by monoubiquitinated histone H3 was additively enhanced by the Uhrf1 SRA domain, which also binds to RFTS. Thus, Dnmt1 activity is regulated by catalysis (ubiquitination)-dependent and -independent functions of Uhrf1.

The position of the vertebral artery V1 segment relative to the C7 vertebra.

BACKGROUND: The vertebral artery (VA) usually enters the transverse foramen at the C6 level. Thus, surgeons prefer to insert pedicle screws (PSs) at C7, but this does not eliminate the risk of VA injury. We aimed to clarify anatomical features of the VA V1 segment at the C7 pedicle level, based on computed tomographic angiography (CTA) of 81 consecutive patients. METHODS: We examined the course of the VA V1 segment on axial CTA images. VA position was classified according to its alignment with the anterior (A), middle (M), or posterior (P) third of the C7 vertebral body at the pedicle level. We also assessed the prevalence of hypoplastic VA (HVA). We measured the distance (VED) from the optimum C7 PS entry point (Ep) to the center of the VA. We also measured the angles formed by the vertebral midline and a line from the inner edge of the VA to the Ep (the VEA), and by the vertebral midline and a line from the inner edge of the pedicle to the Ep (the PEA). RESULTS: The variant location of the VA to the C7 vertebra was A in 13 courses (8.1%), M in 123 (76.9%), and P in 20 (12.5%). HVA was present in the contralateral side in 7 of 20 courses (35%) in the P group, and in 8 of 127 courses (6.3%) in the M group (p < 0.05). The mean VED was 20.2 mm, the mean VEA 6.9°, and the mean PEA angle was 36.3°. CONCLUSION: The 20 VA courses in the P group (12.5% of the total VA courses) were relatively close to the C7 Ep. HVA was present contralateral to the VA in 7 of 20 courses in the P group. CTA should be considered before proceeding with, even if, C7 PS instrumentation, to avoid unexpected pitfall.

Srs2 helicase prevents the formation of toxic DNA damage during late prophase I of yeast meiosis.

Proper repair of double-strand breaks (DSBs) is key to ensure proper chromosome segregation. In this study, we found that the deletion of the SRS2 gene, which encodes a DNA helicase necessary for the control of homologous recombination, induces aberrant chromosome segregation during budding yeast meiosis. This abnormal chromosome segregation in srs2 cells accompanies the formation of a novel DNA damage induced during late meiotic prophase I. The damage may contain long stretches of single-stranded DNAs (ssDNAs), which lead to aggregate formation of a ssDNA binding protein, RPA, and a RecA homolog, Rad51, as well as other recombination proteins inside of the nuclei, but not that of a meiosis-specific Dmc1. The Rad51 aggregate formation in the srs2 mutant depends on the initiation of meiotic recombination and occurs in the absence of chromosome segregation. Importantly, as an early recombination intermediate, we detected a thin bridge of Rad51 between two Rad51 foci in the srs2 mutant, which is rarely seen in wild type. These might be cytological manifestation of the connection of two DSB ends and/or multi-invasion. The DNA damage with Rad51 aggregates in the srs2 mutant is passed through anaphases I and II, suggesting the absence of DNA damage-induced cell cycle arrest after the pachytene stage. We propose that Srs2 helicase resolves early protein-DNA recombination intermediates to suppress the formation of aberrant lethal DNA damage during late prophase I.

Meiosis-specific cohesin component, Rec8, promotes the localization of Mps3 SUN domain protein on the nuclear envelope.

Proteins in the nuclear envelope (NE) play a role in the dynamics and functions of the nucleus and of chromosomes during mitosis and meiosis. Mps3, a yeast NE protein with a conserved SUN domain, predominantly localizes on a yeast centrosome equivalent, spindle pole body (SPB), in mitotic cells. During meiosis, Mps3, together with SPB, forms a distinct multiple ensemble on NE. How meiosis-specific NE localization of Mps3 is regulated remains largely unknown. In this study, we found that a meiosis-specific component of the protein complex essential for sister chromatid cohesion, Rec8, binds to Mps3 during meiosis and controls Mps3 localization and proper dynamics on NE. Ectopic expression of Rec8 in mitotic yeast cells induced the formation of Mps3 patches/foci on NE. This required the cohesin regulator, WAPL ortholog, Rad61/Wpl1, suggesting that a meiosis-specific cohesin complex with Rec8 controls NE localization of Mps3. We also observed that two domains of the nucleoplasmic region of Mps3 are essential for NE localization of Mps3 in mitotic as well as meiotic cells. We speculate that the interaction of Mps3 with the meiosis-specific cohesin in the nucleoplasm is a key determinant for NE localization/function of Mps3.

Influence of novel design alteration of pedicle screw on pull-out strength: A finite element study.

BACKGROUND: We conducted a finite element study to assess the effectiveness of a novel pedicle screw design with two alterations in the distal and proximal portions. METHODS: Finite element (FE) models of 24 vertebrae were constructed using computed tomographic data. Pull-out strength of 4 different pedicle screws were compared. The basic screw design was a dual threaded one (PS0), in which the proximal portion is double-threaded (cortical thread), and the distal portion is single-threaded (cancellous thread). In PS1, the inter-thread double-core shape was added to PS0 in the distal portion. Compared to PS0, in PS2, the proximal portion was elongated by 5 mm. PS3 had both PS1 and PS2 features. In addition, the 24 vertebrae were classified into 3 groups based on volumetric bone mineral density (vBMD) of the vertebral body: low <120 mg/cm3, moderate 120-170 mg/cm3, and high >170 mg/cm3. RESULTS: The mean pull-out strengths (±SD) were 1137 ± 500 N, 1188 ± 520 N, 1191 ± 512 N, and 1242 ± 538 N for PS0, PS1, PS2, and PS3, respectively. In PS1, there was significant difference in the incremental ratio of pull-out strength to PS0 between the low and high vBMD groups (3.7 ± 1.6% vs. 5.0 ± 1.0%, p = 0.006). In PS2, there was a significant difference in the incremental ratio to PS0 between the moderate and high vBMD groups (7.6 ± 4.0% vs. 3.3 ± 1.8%, p < 0.001). In PS3, there was a significant difference in the incremental ratio to PS0 between the moderate and high vBMD groups (12.1 ± 4.8% vs. 8.5 ± 2.1%, p = 0.003). CONCLUSIONS: The two design alterations showed the combined additive effect in the PS3 design. The moderate vBMD group has a balanced bone property to reflect the combined effects of the PS1 and PS2 design alterations.

Genetic Interactions of Histone Modification Machinery Set1 and PAF1C with the Recombination Complex Rec114-Mer2-Mei4 in the Formation of Meiotic DNA Double-Strand Breaks.

Homologous recombination is essential for chromosome segregation during meiosis I. Meiotic recombination is initiated by the introduction of double-strand breaks (DSBs) at specific genomic locations called hotspots, which are catalyzed by Spo11 and its partners. DSB hotspots during meiosis are marked with Set1-mediated histone H3K4 methylation. The Spo11 partner complex, Rec114-Mer2-Mei4, essential for the DSB formation, localizes to the chromosome axes. For efficient DSB formation, a hotspot with histone H3K4 methylation on the chromatin loops is tethered to the chromosome axis through the H3K4 methylation reader protein, Spp1, on the axes, which interacts with Mer2. In this study, we found genetic interaction of mutants in a histone modification protein complex called PAF1C with the REC114 and MER2 in the DSB formation in budding yeast Saccharomyces cerevisiae. Namely, the paf1c mutations rtf1 and cdc73 showed synthetic defects in meiotic DSB formation only when combined with a wild-type-like tagged allele of either the REC114 or MER2. The synthetic defect of the tagged REC114 allele in the DSB formation was seen also with the set1, but not with spp1 deletion. These results suggest a novel role of histone modification machinery in DSB formation during meiosis, which is independent of Spp1-mediated loop-axis tethering.