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1.
J C Game 《Developmental genetics》1992,13(6):485-497
Pulsed-field gel electrophoresis (PFGE) has been used to study the timing, frequency, and distribution of double-strand breaks (DSBs) in chromosomal-sized DNA during meiosis in yeast. It has previously been shown that DSBs are associated with some genetic hotspots during recombination, and it is important to know whether meiotic recombination events routinely initiate via DSBs. Two strains have been studied here--a high-sporulating homothallic wild type and a congenic mutant strain carrying a rad50S mutation. This mutant has previously been reported to accumulate broken molecules in meiosis to much higher frequencies than wild type and to abolish the characteristic wild-type processing of DNA that has been observed at the break sites. When whole chromosomes are resolved by PFGE, both strains show some broken molecules starting at the time that cells commit to genetic recombination. Breakage has been assessed primarily on Chromosome III and Chr. XV, using Southern hybridization to identify these chromosomes and their fragments. At any one time, break frequency in wild type is much lower than the cumulative frequency of recombination events that occur during meiosis. However, there is suggestive evidence that each break is short-lived, and it is therefore difficult to estimate the total number of breaks that may occur. In rad50S, chromosome breaks accumulate to much higher levels, which are probably broadly consistent with the estimated number of recombination events in wild type. However, since rad50S is substantially defective in completing recombination, it is not known for certain if it initiates events at wild-type frequencies. A surprising feature of the data is that a strong banding pattern is observed in the fragment distribution from broken chromosomes in both strains, implying that at least much of the breakage occurs at specific sites or within short regions. However, with the exception of the rDNA region on Chr. XII, assessment of the genetic map indicates that recombination can occur almost anywhere in the genome, although some regions are much hotter than others. Possible reasons for this apparent paradox are discussed. It may in part result from breakage levels too low for adequate detection in cold regions but may also imply that recombination events are localized more than previously realized. Alternatively, there may be a more indirect relationship between break sites and the associated recombination events. 相似文献
2.
Meiotic Recombination Initiated by a Double-Strand Break in Rad50Δ Yeast Cells Otherwise Unable to Initiate Meiotic Recombination
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Meiotic recombination in Saccharomyces cerevisiae is initiated by double-strand breaks (DSBs). We have developed a system to compare the properties of meiotic DSBs with those created by the site-specific HO endonuclease. HO endonuclease was expressed under the control of the meiotic-specific SPO13 promoter, creating a DSB at a single site on one of yeast's 16 chromosomes. In Rad(+) strains the times of appearance of the HO-induced DSBs and of subsequent recombinants are coincident with those induced by normal meiotic DSBs. Physical monitoring of DNA showed that SPO13::HO induced gene conversions both in Rad(+) and in rad50Δ cells that cannot initiate normal meiotic DSBs. We find that the RAD50 gene is important, but not essential, for recombination even after a DSB has been created in a meiotic cell. In rad50Δ cells, some DSBs are not repaired until a broken chromosome has been packaged into a spore and is subsequently germinated. This suggests that a broken chromosome does not signal an arrest of progression through meiosis. The recombination defect in rad50Δ diploids is not, however, meiotic specific, as mitotic rad50 diploids, experiencing an HO-induced DSB, exhibit similar departures from wild-type recombination. 相似文献
3.
The fission yeast Schizosaccharomyces pombe Rec12 protein, the homolog of Spo11 in other organisms, initiates meiotic recombination by creating DNA double-strand breaks (DSBs) and becoming covalently linked to the DNA ends of the break. This protein–DNA linkage has previously been detected only in mutants such as rad50S in which break repair is impeded and DSBs accumulate. In the budding yeast Saccharomyces cerevisiae, the DSB distribution in a rad50S mutant is markedly different from that in wild-type (RAD50) meiosis, and it was suggested that this might also be true for other organisms. Here, we show that we can detect Rec12-DNA linkages in Sc. pombe rad50+ cells, which are proficient for DSB repair. In contrast to the results from Sa. cerevisiae, genome-wide microarray analysis of Rec12-DNA reveals indistinguishable meiotic DSB distributions in rad50+ and rad50S strains of Sc. pombe. These results confirm our earlier findings describing the occurrence of widely spaced DSBs primarily in large intergenic regions of DNA and demonstrate the relevance and usefulness of fission yeast studies employing rad50S. We propose that the differential behavior of rad50S strains reflects a major difference in DSB regulation between the two species—specifically, the requirement for the Rad50-containing complex for DSB formation in budding yeast but not in fission yeast. Use of rad50S and related mutations may be a useful method for DSB analysis in other species. 相似文献
4.
Mapping Meiotic Single-Strand DNA Reveals a New Landscape of DNA Double-Strand Breaks in Saccharomyces cerevisiae 总被引:1,自引:0,他引:1
DNA double-strand breaks (DSBs), which are formed by the Spo11 protein, initiate meiotic recombination. Previous DSB-mapping studies have used rad50S or sae2Δ mutants, which are defective in break processing, to accumulate Spo11-linked DSBs, and report large (≥ 50 kb) “DSB-hot” regions that are separated by “DSB-cold” domains of similar size. Substantial recombination occurs in some DSB-cold regions, suggesting that DSB patterns are not normal in rad50S or sae2Δ mutants. We therefore developed a novel method to map genome-wide, single-strand DNA (ssDNA)–associated DSBs that accumulate in processing-capable, repair-defective dmc1 Δ and dmc1 Δ rad51 Δ mutants. DSBs were observed at known hot spots, but also in most previously identified “DSB-cold” regions, including near centromeres and telomeres. Although approximately 40% of the genome is DSB-cold in rad50S mutants, analysis of meiotic ssDNA from dmc1Δ shows that most of these regions have substantial DSB activity. Southern blot assays of DSBs in selected regions in dmc1Δ , rad50S , and wild-type cells confirm these findings. Thus, DSBs are distributed much more uniformly than was previously believed. Comparisons of DSB signals in dmc1 , dmc1 rad51 , and dmc1 spo11 mutant strains identify Dmc1 as a critical strand-exchange activity genome-wide, and confirm previous conclusions that Spo11-induced lesions initiate all meiotic recombination. 相似文献
5.
Mnd1/Hop2 facilitates Dmc1-dependent interhomolog crossover formation in meiosis of budding yeast
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Henry JM Camahort R Rice DA Florens L Swanson SK Washburn MP Gerton JL 《Molecular and cellular biology》2006,26(8):2913-2923
During meiosis, each chromosome must pair with its homolog and undergo meiotic crossover recombination in order to segregate properly at the first meiotic division. Recombination in meiosis in Saccharomyces cerevisiae relies on two Escherichia coli recA homologs, Rad51 and Dmc1, as well as the more recently discovered heterodimer Mnd1/Hop2. Meiotic recombination in S. cerevisiae mnd1 and hop2 single mutants is initiated via double-strand breaks (DSBs) but does not progress beyond this stage; heteroduplex DNA, joint molecules, and crossovers are not detected. Whereas hop2 and mnd1 single mutants are profoundly recombination defective, we show that mnd1 rad51, hop2 rad51, and mnd1 rad17 double mutants are able to carry out crossover recombination. Interestingly, noncrossover recombination is absent, indicating a role for Mnd1/Hop2 in the designation of DSBs for noncrossover recombination. We demonstrate that in the rad51 mnd1 double mutant, recombination is more likely to occur between repetitive sequences on nonhomologous chromosomes. Our results support a model in which Mnd1/Hop2 is required for DNA-DNA interactions that help ensure Dmc1-mediated stable strand invasion between homologous chromosomes, thereby preserving genomic integrity. 相似文献
6.
Multiple sites for double-strand breaks in whole meiotic chromosomes of Saccharomyces cerevisiae. 总被引:16,自引:4,他引:12
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We present a scheme for locating double-strand breaks (DSBs) in meiotic chromosomes of Saccharomyces cerevisiae, based on the separation of large DNA molecules by pulsed field gel electrophoresis. Using a rad50S mutant, in which DSBs are not processed, we show that DSBs are widely induced in S. cerevisiae chromosomes during meiosis. Some of the DSBs accumulate at certain preferred sites. We present general profiles of DSBs in chromosomes III, V, VI and VII. A map of the 12 preferred sites on chromosome III is presented. At least some of these sites correlate with known 'hot spots' for meiotic recombination. The data are discussed in view of current models of meiotic recombination and chromosome segregation. 相似文献
7.
DNA double-strand breaks (DSBs), which are formed by the Spo11 protein, initiate meiotic recombination. Previous DSB-mapping studies have used rad50S or sae2Δ mutants, which are defective in break processing, to accumulate Spo11-linked DSBs, and report large (≥ 50 kb) “DSB-hot” regions that are separated by “DSB-cold” domains of similar size. Substantial recombination occurs in some DSB-cold regions, suggesting that DSB patterns are not normal in rad50S or sae2Δ mutants. We therefore developed a novel method to map genome-wide, single-strand DNA (ssDNA)–associated DSBs that accumulate in processing-capable, repair-defective dmc1Δ and dmc1Δ rad51Δ mutants. DSBs were observed at known hot spots, but also in most previously identified “DSB-cold” regions, including near centromeres and telomeres. Although approximately 40% of the genome is DSB-cold in rad50S mutants, analysis of meiotic ssDNA from dmc1Δ shows that most of these regions have substantial DSB activity. Southern blot assays of DSBs in selected regions in dmc1Δ, rad50S, and wild-type cells confirm these findings. Thus, DSBs are distributed much more uniformly than was previously believed. Comparisons of DSB signals in dmc1, dmc1 rad51, and dmc1 spo11 mutant strains identify Dmc1 as a critical strand-exchange activity genome-wide, and confirm previous conclusions that Spo11-induced lesions initiate all meiotic recombination. 相似文献
8.
Involvement of Sir2/4 in silencing of DNA breakage and recombination on mouse YACs during yeast meiosis
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Klieger Y Yizhar O Zenvirth D Shtepel-Milman N Snoek M Simchen G 《Molecular biology of the cell》2005,16(3):1449-1455
Yeast artificial chromosomes (YACs) that contain human DNA backbone undergo DNA double-strand breaks (DSBs) and recombination during yeast meiosis at rates similar to the yeast native chromosomes. Surprisingly, YACs containing DNA covering a recombination hot spot in the mouse major histocompatibility complex class III region do not show meiotic DSBs and undergo meiotic recombination at reduced levels. Moreover, segregation of these YACs during meiosis is seriously compromised. In meiotic yeast cells carrying the mutations sir2 or sir4, but not sir3, these YACs show DSBs, suggesting that a unique chromatin structure of the YACs, involving Sir2 and Sir4, protects the YACs from the meiotic recombination machinery. We speculate that the paucity of DSBs and recombination events on these YACs during yeast meiosis may reflect the refractory nature of the corresponding region in the mouse genome. 相似文献
9.
Li J Harper LC Golubovskaya I Wang CR Weber D Meeley RB McElver J Bowen B Cande WZ Schnable PS 《Genetics》2007,176(3):1469-1482
In Saccharomyces cerevisiae, Rad51p plays a central role in homologous recombination and the repair of double-strand breaks (DSBs). Double mutants of the two Zea mays L. (maize) rad51 homologs are viable and develop well under normal conditions, but are male sterile and have substantially reduced seed set. Light microscopic analyses of male meiosis in these plants reveal reduced homologous pairing, synapsis of nonhomologous chromosomes, reduced bivalents at diakinesis, numerous chromosome breaks at anaphase I, and that >33% of quartets carry cells that either lack an organized nucleolus or have two nucleoli. This indicates that RAD51 is required for efficient chromosome pairing and its absence results in nonhomologous pairing and synapsis. These phenotypes differ from those of an Arabidopsis rad51 mutant that exhibits completely disrupted chromosome pairing and synapsis during meiosis. Unexpectedly, surviving female gametes produced by maize rad51 double mutants are euploid and exhibit near-normal rates of meiotic crossovers. The finding that maize rad51 double mutant embryos are extremely susceptible to radiation-induced DSBs demonstrates a conserved role for RAD51 in the repair of mitotic DSBs in plants, vertebrates, and yeast. 相似文献
10.
Recombination at subtelomeres is regulated by physical distance,double‐strand break resection and chromatin status
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Hélène Bordelet Antoine Hocher Myriam Ruault Adouda Adjiri Karine Dubrana 《The EMBO journal》2017,36(17):2609-2625
Homologous recombination (HR) is a conserved mechanism that repairs broken chromosomes via intact homologous sequences. How different genomic, chromatin and subnuclear contexts influence HR efficiency and outcome is poorly understood. We developed an assay to assess HR outcome by gene conversion (GC) and break‐induced replication (BIR), and discovered that subtelomeric double‐stranded breaks (DSBs) are preferentially repaired by BIR despite the presence of flanking homologous sequences. Overexpression of a silencing‐deficient SIR3 mutant led to active grouping of telomeres and specifically increased the GC efficiency between subtelomeres. Thus, physical distance limits GC at subtelomeres. However, the repair efficiency between reciprocal intrachromosomal and subtelomeric sequences varies up to 15‐fold, depending on the location of the DSB, indicating that spatial proximity is not the only limiting factor for HR. EXO1 deletion limited the resection at subtelomeric DSBs and improved GC efficiency. The presence of repressive chromatin at subtelomeric DSBs also favoured recombination, by counteracting EXO1‐mediated resection. Thus, repressive chromatin promotes HR at subtelomeric DSBs by limiting DSB resection and protecting against genetic information loss. 相似文献
11.
Rice RAD51 paralogs play essential roles in somatic homologous recombination for DNA repair
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Zhan Xu Jianxiang Zhang Meng Xu Wen Ji Meimei Yu Yajun Tao Zhiyun Gong Minghong Gu Hengxiu Yu 《The Plant journal : for cell and molecular biology》2018,95(2):282-295
Synthesis‐dependent strand annealing (SDSA) and single‐strand annealing (SSA) are the two main homologous recombination (HR) pathways in double‐strand break (DSB) repair. The involvement of rice RAD51 paralogs in HR is well known in meiosis, although the molecular mechanism in somatic HR remains obscure. Loss‐of‐function mutants of rad51 paralogs show increased sensitivity to the DSB‐inducer bleomycin, which results in greatly compromised somatic recombination efficiencies (xrcc3 in SDSA, rad51b and xrcc2 in SSA, rad51c and rad51d in both). Using immunostaining, we found that mutations in RAD51 paralogs (XRCC3, RAD51C, or RAD51D) lead to tremendous impairment in RAD51 focus formation at DSBs. Intriguingly, the RAD51C mutation has a strong effect on the protein loading of its partners (XRCC3 and RAD51B) at DSBs, which is similar to the phenomenon observed in the case of blocking PI3K‐like kinases in wild‐type plant. We conclude that the rice CDX3 complex acts in SDSA recombination while the BCDX2 complex acts in SSA recombination in somatic DSB repair. Importantly, RAD51C serves as a fulcrum for the local recruitment of its partners (XRCC3 for SDSA and RAD51B for SSA) and is positively modulated by PI3K‐like kinases to facilitate both the SDSA and SSA pathways in RAD51 paralog‐dependent somatic HR. 相似文献
12.
In Saccharomyces cerevisiae, meiotic recombination is initiated by transient DNA double-stranded breaks (DSBs). These DSBs undergo a 5' --> 3' resection to produce 3' single-stranded DNA ends that serve to channel DSBs into the RAD52 recombinational repair pathway. In vitro studies strongly suggest that several proteins of this pathway--Rad51, Rad52, Rad54, Rad55, Rad57, and replication protein A (RPA)--play a role in the strand exchange reaction. Here, we report a study of the meiotic phenotypes conferred by two missense mutations affecting the largest subunit of RPA, which are localized in the protein interaction domain (rfa1-t11) and in the DNA-binding domain (rfa1-t48). We find that both mutant diploids exhibit reduced sporulation efficiency, very poor spore viability, and a 10- to 100-fold decrease in meiotic recombination. Physical analyses indicate that both mutants form normal levels of meiosis-specific DSBs and that the broken ends are processed into 3'-OH single-stranded tails, indicating that the RPA complex present in these rfa1 mutants is functional in the initial steps of meiotic recombination. However, the 5' ends of the broken fragments undergo extensive resection, similar to what is observed in rad51, rad52, rad55, and rad57 mutants, indicating that these RPA mutants are defective in the repair of the Spo11-dependent DSBs that initiate homologous recombination during meiosis. 相似文献
13.
Yang Liu Yingtian Deng Gang Li Jie Zhao 《The Plant journal : for cell and molecular biology》2013,73(1):154-165
Replication factor C1 (RFC1), which is conserved in eukaryotes, is involved in DNA replication and checkpoint control. However, a RFC1 product participating in DNA repair at meiosis has not been reported in Arabidopsis. Here, we report functional characterization of AtRFC1 through analysis of the rfc1–2 mutant. The rfc1–2 mutant displayed normal vegetative growth but showed silique sterility because the male gametophyte was arrested at the uninucleus microspore stage and the female at the functional megaspore stage. Expression of AtRFC1 was concentrated in the reproductive organ primordia, meiocytes and developing gametes. Chromosome spreads showed that pairing and synapsis were normal, and the chromosomes were broken when desynapsis began at late prophase I, and chromosome fragments remained in the subsequent stages. For this reason, homologous chromosomes and sister chromatids segregated unequally, leading to pollen sterility. Immunolocalization revealed that the AtRFC1 protein localized to the chromosomes during zygotene and pachytene in wild‐type but were absent in the spo11–1 mutant. The chromosome fragmentation of rfc1–2 was suppressed by spo11–1, indicating that AtRFC1 acted downstream of AtSPO11‐1. The similar chromosome behavior of rad51 rfc1–2 and rad51 suggests that AtRFC1 may act with AtRAD51 in the same pathway. In summary, AtRFC1 is required for DNA double‐strand break repair during meiotic homologous recombination of Arabidopsis. 相似文献
14.
Characterization of the Roles of the Saccharomyces Cerevisiae Rad54 Gene and a Homologue of Rad54, Rdh54/Tid1, in Mitosis and Meiosis 总被引:4,自引:0,他引:4
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M. Shinohara E. Shita-Yamaguchi J. M. Buerstedde H. Shinagawa H. Ogawa A. Shinohara 《Genetics》1997,147(4):1545-1556
The RAD54 gene, which encodes a protein in the SWI2/SNF2 family, plays an important role in recombination and DNA repair in Saccharomyces cerevisiae. The yeast genome project revealed a homologue of RAD54, RDH54/TID1. Properties of the rdh54/tid1 mutant and the rad54 rdh54/tid1 double mutant are shown for mitosis and meiosis. The rad54 mutant is sensitive to the alkylating agent, methyl methanesulfonate (MMS), and is defective in interchromosomal and intrachromosomal gene conversion. The rdh54/tid1 single mutant, on the other hand, does not show any significant deficiency in mitosis. However, the rad54 rdh54/tid1 mutant is more sensitive to MMS and more defective in interchromosomal gene conversion than is the rad54 mutant, but shows the same frequency of intrachromosomal gene conversion as the rad54 mutant. These results suggest that RDH54/TID1 is involved in a minor pathway of mitotic recombination in the absence of RAD54. In meiosis, both single mutants produce viable spores at slightly reduced frequency. However, only the rdh54/tid1 mutant, but not the rad54 mutant, shows significant defects in recombination: retardation of the repair of meiosis-specific double-strand breaks (DSBs) and delayed formation of physical recombinants. Furthermore, the rad54 rdh54/tid1 double mutant is completely defective in meiosis, accumulating DSBs with more recessed ends than the wild type and producing fewer physical recombinants than the wild type. These results suggest that one of the differences between the late stages of mitotic recombination and meiotic recombination might be specified by differential dependency on the Rad54 and Rdh54/Tid1 proteins. 相似文献
15.
The location and structure of double-strand DNA breaks induced during yeast meiosis: evidence for a covalently linked DNA-protein intermediate. 总被引:19,自引:2,他引:17
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We have determined the precise location and structure of the double-strand DNA breaks (DSBs) formed during Saccharomyces cerevisiae meiosis. Breaks were examined at two recombination hot spots in both wild-type and rad50S mutant cells. At both loci, breaks occurred at multiple, irregularly spaced sites in a approximately 150 nucleotide interval contained within an area of nuclease-hypersensitive chromatin. No consensus sequence could be discerned at or around break sites. Patterns of cleavage observed on individual strands indicated that breaks initially form with a two nucleotide 5' overhang. Broken strands from rad50S mutant cells contained tightly bound protein at their 5' ends. We suggest that, in S.cerevisiae, meiotic recombination is initiated by a DSB-forming activity that creates a covalently linked protein-DNA intermediate. 相似文献
16.
《Fly》2013,7(2):134-140
During prophase of meiosis I, genetic recombination is initiated with a Spo11-dependent DNA double-strand break (DSB). Repair of these DSBs can generate crossovers, which become chiasmata and are important for the process of chromosome segregation. To ensure at least one chiasma per homologous pair of chromosomes, the number and distribution of crossovers is regulated. One system contributing to the distribution of crossovers is the pachytene checkpoint, which requires the conserved gene pch2 that encodes an AAA+ATPase family member. Pch2-dependent pachytene checkpoint function causes delays in pachytene progression when there are defects in processes required for crossover formation, such as mutations in DSB-repair genes and when there are defects in the structure of the meiotic chromosome axis. Thus, the pachytene checkpoint appears to monitor events leading up to the generation of crossovers. Interestingly, heterozygous chromosome rearrangements cause Pch2-dependent pachytene delays and as little as two breaks in the continuity of the paired chromosome axes are sufficient to evoke checkpoint activity. These chromosome rearrangements also cause an interchromosomal effect on recombination whereby crossing over is suppressed between the affected chromosomes but is increased between the normal chromosome pairs. We have shown that this phenomenon is also due to pachytene checkpoint activity. 相似文献
17.
Jeremy S. Bickel Liting Chen Jin Hayward Szu Ling Yeap Ashley E. Alkers Raymond C. Chan 《PLoS genetics》2010,6(7)
In meiosis, programmed DNA breaks repaired by homologous recombination (HR) can be processed into inter-homolog crossovers that promote the accurate segregation of chromosomes. In general, more programmed DNA double-strand breaks (DSBs) are formed than the number of inter-homolog crossovers, and the excess DSBs must be repaired to maintain genomic stability. Sister-chromatid (inter-sister) recombination is postulated to be important for the completion of meiotic DSB repair. However, this hypothesis is difficult to test because of limited experimental means to disrupt inter-sister and not inter-homolog HR in meiosis. We find that the conserved Structural Maintenance of Chromosomes (SMC) 5 and 6 proteins in Caenorhabditis elegans are required for the successful completion of meiotic homologous recombination repair, yet they appeared to be dispensable for accurate chromosome segregation in meiosis. Mutations in the smc-5 and smc-6 genes induced chromosome fragments and dismorphology. Chromosome fragments associated with HR defects have only been reported in mutants, which have disrupted inter-homolog crossover. Surprisingly, the smc-5 and smc-6 mutations did not disrupt the formation of chiasmata, the cytologically visible linkages between homologous chromosomes formed from meiotic inter-homolog crossovers. The mutant fragmentation defect appeared to be preferentially enhanced by the disruptions of inter-homolog recombination but not by the disruptions of inter-sister recombination. Based on these findings, we propose that the C. elegans SMC-5/6 proteins are required in meiosis for the processing of homolog-independent, presumably sister-chromatid-mediated, recombination repair. Together, these results demonstrate that the successful completion of homolog-independent recombination is crucial for germ cell genomic stability. 相似文献
18.
Temporal comparison of recombination and synaptonemal complex formation during meiosis in S. cerevisiae 总被引:53,自引:0,他引:53
In synchronous cultures of S. cerevisiae undergoing meiosis, an early event in the meiotic recombination pathway, site-specific double strand breaks (DSBs), occurs early in prophase, in some instances well before tripartite synaptonemal complex (SC) begins to form. This observation, together with previous results, supports the view that events involving DSBs are required for SC formation. We discuss the possibility that the mitotic pathway for recombinational repair of DSBs served as the primordial mechanism for connecting homologous chromosomes during the evolution of meiosis. DSBs disappear during the period when tripartite SC structure is forming and elongating (zygotene); presumably, they are converted to another type of recombination intermediate. Neither DSBs nor mature recombinant molecules are present when SCs are full length (pachytene). Mature reciprocally recombinant molecules arise at the end of or just after pachytene. We suggest that the SC might coordinate recombinant maturation with other events of meiosis. 相似文献
19.
Marie-Therese Kurzbauer Michael Peter Janisiw Luis F Paulin Ignacio Prusn Mota Konstantin Tomanov Ondrej Krsicka Arndt von Haeseler Veit Schubert Peter Schlgelhofer 《The Plant cell》2021,33(5):1633
Meiosis is a specialized cell division that gives rise to genetically distinct gametic cells. Meiosis relies on the tightly controlled formation of DNA double-strand breaks (DSBs) and their repair via homologous recombination for correct chromosome segregation. Like all forms of DNA damage, meiotic DSBs are potentially harmful and their formation activates an elaborate response to inhibit excessive DNA break formation and ensure successful repair. Previous studies established the protein kinase ATM as a DSB sensor and meiotic regulator in several organisms. Here we show that Arabidopsis ATM acts at multiple steps during DSB formation and processing, as well as crossover (CO) formation and synaptonemal complex (SC) organization, all vital for the successful completion of meiosis. We developed a single-molecule approach to quantify meiotic breaks and determined that ATM is essential to limit the number of meiotic DSBs. Local and genome-wide recombination screens showed that ATM restricts the number of interference-insensitive COs, while super-resolution STED nanoscopy of meiotic chromosomes revealed that the kinase affects chromatin loop size and SC length and width. Our study extends our understanding of how ATM functions during plant meiosis and establishes it as an integral factor of the meiotic program.Arabidopsis ATM acts at multiple steps during DSB formation and processing, as well as crossover formation and synaptonemal complex organization, all vital for the successful completion of meiosis. 相似文献
20.
An Implanted Recombination Hot Spot Stimulates Recombination and Enhances Sister Chromatid Cohesion of Heterologous Yacs during Yeast Meiosis 总被引:2,自引:0,他引:2
Heterologous yeast artificial chromosomes (YACs) do not recombine with each other and missegregate in 25% of meiosis I events. Recombination hot spots in the yeast Saccharomyces cerevisiae have previously been shown to be associated with sites of meiosis-induced double-strand breaks (DSBs). A 6-kb fragment containing a recombination hot spot/DSB site was implanted onto two heterologous human DNA YACs and was shown to cause the YACs to undergo meiotic recombination in 5-8% of tetrads. Reciprocal exchanges initiated and resolved within the 6-kb insert. Presence of the insert had no detectable effect on meiosis I nondisjunction. Surprisingly, the recombination hot spots acted in cis to significantly reduce precocious sister-chromatid segregation. This novel observation suggests that DSBs are instrumental in maintaining cohesion between sister chromatids in meiosis I. We propose that this previously unknown function of DSBs is mediated by the stimulation of sister-chromatid exchange and/or its intermediates. 相似文献