Coordination of meiotic recombination, pairing, and synapsis by PHS1

Pairing, synapsis, and recombination are prerequisites for accurate chromosome segregation in meiosis. The phs1 gene in maize is required for pairing to occur between homologous chromosomes. In the phs1 mutant, homologous chromosome synapsis is completely replaced by synapsis between nonhomologous partners. The phs1 gene is also required for installation of the meiotic recombination machinery on chromosomes, as the mutant almost completely lacks chromosomal foci of the recombination protein RAD51. Thus, in the phs1 mutant, synapsis is uncoupled from recombination and pairing. The protein encoded by the phs1 gene likely acts in a multistep process to coordinate pairing, recombination, and synapsis.     

Mechanism of RAD51-dependent DNA interstrand cross-link repair

DNA interstrand cross-links (ICLs) are toxic DNA lesions whose repair in S phase of eukaryotic cells is incompletely understood. In Xenopus egg extracts, ICL repair is initiated when two replication forks converge on the lesion. Dual incisions then create a DNA double-strand break (DSB) in one sister chromatid, whereas lesion bypass restores the other sister. We report that the broken sister chromatid is repaired via RAD51-dependent strand invasion into the regenerated sister. Recombination acts downstream of FANCI-FANCD2, yet RAD51 binds ICL-stalled replication forks independently of FANCI-FANCD2 and before DSB formation. Our results elucidate the functional link between the Fanconi anemia pathway and the recombination machinery during ICL repair. In addition, they demonstrate the complete repair of a DSB via homologous recombination in vitro.     

BRCA2 function in DNA binding and recombination from a BRCA2-DSS1-ssDNA structure

Mutations in the BRCA2 (breast cancer susceptibility gene 2) tumor suppressor lead to chromosomal instability due to defects in the repair of double-strand DNA breaks (DSBs) by homologous recombination, but BRCA2's role in this process has been unclear. Here, we present the 3.1 angstrom crystal structure of a approximately 90-kilodalton BRCA2 domain bound to DSS1, which reveals three oligonucleotide-binding (OB) folds and a helix-turn-helix (HTH) motif. We also (i) demonstrate that this BRCA2 domain binds single-stranded DNA, (ii) present its 3.5 angstrom structure bound to oligo(dT)9, (iii) provide data that implicate the HTH motif in dsDNA binding, and (iv) show that BRCA2 stimulates RAD51-mediated recombination in vitro. These findings establish that BRCA2 functions directly in homologous recombination and provide a structural and biochemical basis for understanding the loss of recombination-mediated DSB repair in BRCA2-associated cancers.     

RAD51C is required for Holliday junction processing in mammalian cells

During genetic recombination and the recombinational repair of chromosome breaks, DNA molecules become linked at points of strand exchange. Branch migration and resolution of these crossovers, or Holliday junctions (HJs), complete the recombination process. Here, we show that extracts from cells carrying mutations in the recombination/repair genes RAD51C or XRCC3 have reduced levels of HJ resolvase activity. Moreover, depletion of RAD51C from fractionated human extracts caused a loss of branch migration and resolution activity, but these functions were restored by complementation with a variety of RAD51 paralog complexes containing RAD51C. We conclude that the RAD51 paralogs are involved in HJ processing in human cells.     

A conserved checkpoint monitors meiotic chromosome synapsis in Caenorhabditis elegans

We report the discovery of a checkpoint that monitors synapsis between homologous chromosomes to ensure accurate meiotic segregation. Oocytes containing unsynapsed chromosomes selectively undergo apoptosis even if a germline DNA damage checkpoint is inactivated. This culling mechanism is specifically activated by unsynapsed pairing centers, cis-acting chromosome sites that are also required to promote synapsis in Caenorhabditis elegans. Apoptosis due to synaptic failure also requires the C. elegans homolog of PCH2, a budding yeast pachytene checkpoint gene, which suggests that this surveillance mechanism is widely conserved.     

小麦PH基因系的研究及应用

小麦中存在着抑制部分同源染色体配对的两个主效基因ＰＨ１和ＰＨ２。此外,还携带有许多微效的抑制或促进配对基因。小麦染色体配对过程是受一个多基因系统控制,相当复杂。ＰＨ基因制约减数分裂前当染色体的联会。     

Essential role of Fkbp6 in male fertility and homologous chromosome pairing in meiosis

Meiosis is a critical stage of gametogenesis in which alignment and synapsis of chromosomal pairs occur, allowing for the recombination of maternal and paternal genomes. Here we show that FK506 binding protein (Fkbp6) localizes to meiotic chromosome cores and regions of homologous chromosome synapsis. Targeted inactivation of Fkbp6 in mice results in aspermic males and the absence of normal pachytene spermatocytes. Moreover, we identified the deletion of Fkbp6 exon 8 as the causative mutation in spontaneously male sterile as/as mutant rats. Loss of Fkbp6 results in abnormal pairing and misalignments between homologous chromosomes, nonhomologous partner switches, and autosynapsis of X chromosome cores in meiotic spermatocytes. Fertility and meiosis are normal in Fkbp6 mutant females. Thus, Fkbp6 is a component of the synaptonemal complex essential for sex-specific fertility and for the fidelity of homologous chromosome pairing in meiosis.     

Robust crossover assurance and regulated interhomolog access maintain meiotic crossover number

Most organisms rely on interhomolog crossovers (COs) to ensure proper meiotic chromosome segregation but make few COs per chromosome pair. By monitoring repair events at a defined double-strand break (DSB) site during Caenorhabditis elegans meiosis, we reveal mechanisms that ensure formation of the obligate CO while limiting CO number. We find that CO is the preferred DSB repair outcome in the absence of inhibitory effects of other (nascent) recombination events. Thus, a single DSB per chromosome pair is largely sufficient to ensure CO formation. Further, we show that access to the homolog as a repair template is regulated, shutting down simultaneously for both CO and noncrossover (NCO) pathways. We propose that regulation of interhomolog access limits CO number and contributes to CO interference.     

Chromosome choreography: the meiotic ballet

The separation of homologous chromosomes during meiosis in eukaryotes is the physical basis of Mendelian inheritance. The core of the meiotic process is a specialized nuclear division (meiosis I) in which homologs pair with each other, recombine, and then segregate from each other. The processes of chromosome alignment and pairing allow for homolog recognition. Reciprocal meiotic recombination ensures meiotic chromosome segregation by converting sister chromatid cohesion into mechanisms that hold homologous chromosomes together. Finally, the ability of sister kinetochores to orient to a single pole at metaphase I allows the separation of homologs to two different daughter cells. Failures to properly accomplish this elegant chromosome dance result in aneuploidy, a major cause of miscarriage and birth defects in human beings.     

SIRT6 promotes DNA repair under stress by activating PARP1

Sirtuin 6 (SIRT6) is a mammalian homolog of the yeast Sir2 deacetylase. Mice deficient for SIRT6 exhibit genome instability. Here, we show that in mammalian cells subjected to oxidative stress SIRT6 is recruited to the sites of DNA double-strand breaks (DSBs) and stimulates DSB repair, through both nonhomologous end joining and homologous recombination. Our results indicate that SIRT6 physically associates with poly[adenosine diphosphate (ADP)-ribose] polymerase 1 (PARP1) and mono-ADP-ribosylates PARP1 on lysine residue 521, thereby stimulating PARP1 poly-ADP-ribosylase activity and enhancing DSB repair under oxidative stress.     

The fission yeast FANCM ortholog directs non-crossover recombination during meiosis

The formation of healthy gametes depends on programmed DNA double-strand breaks (DSBs), which are each repaired as a crossover (CO) or non-crossover (NCO) from a homologous template. Although most of these DSBs are repaired without giving COs, little is known about the genetic requirements of NCO-specific recombination. We show that Fml1, the Fanconi anemia complementation group M (FANCM)-ortholog of Schizosaccharomyces pombe, directs the formation of NCOs during meiosis in competition with the Mus81-dependent pro-CO pathway. We also define the Rad51/Dmc1-mediator Swi5-Sfr1 as a major determinant in biasing the recombination process in favor of Mus81, to ensure the appropriate amount of COs to guide meiotic chromosome segregation. The conservation of these proteins from yeast to humans suggests that this interplay may be a general feature of meiotic recombination.     

硬粒小麦-簇毛麦双二倍体与普通小麦杂种F1的花粉母细胞减数分裂行为的研究

用普通小麦与硬粒小麦 -簇毛麦双二倍体杂交 ,对其杂种F1的花粉母细胞中期Ⅰ的染色体配对进行了洋红染色和Giemsa -C带染色观察。结果表明虽然簇毛麦染色体在中期Ⅰ主要以单价体存在 ,但也有少量的小麦 -簇毛染色体配对 (平均每细胞为 1 1%左右 ) ,可以通过遗传重组转移簇毛麦的有利基因。这些小麦 -簇毛麦染色体配对 ,既分布在超过理论配对数的细胞中 ,也存在于少于理论配对数的细胞中 ;相反 ,超过理论配对数的染色体对不一定就是小麦 -簇毛麦染色体之间的配对 ,在少于理论配对数的细胞中 ,也不一定仅是小麦同源染色体的配对 ,同样包含有少量的小麦 -簇毛麦染色体配对。因而 ,用传统的染色法对外源物种与小麦染色体之间的配对数的估计 ,既有夸大一部分信息 ,又有掩盖一些有益信息的弊端。说明我们在用传统方法来推测远缘杂种后代中部分同源染色体配对时要持审慎的态度 ,需要用较为准确的方法来直接鉴定     

Rad51 is an accessory factor for Dmc1-mediated joint molecule formation during meiosis

Meiotic recombination in budding yeast requires two RecA-related proteins, Rad51 and Dmc1, both of which form filaments on DNA capable of directing homology search and catalyzing formation of homologous joint molecules (JMs) and strand exchange. With use of a separation-of-function mutant form of Rad51 that retains filament-forming but not JM-forming activity, we show that the JM activity of Rad51 is fully dispensable for meiotic recombination. The corresponding mutation in Dmc1 causes a profound recombination defect, demonstrating Dmc1's JM activity alone is responsible for meiotic recombination. We further provide biochemical evidence that Rad51 acts with Mei5-Sae3 as a Dmc1 accessory factor. Thus, Rad51 is a multifunctional protein that catalyzes recombination directly in mitosis and indirectly, via Dmc1, during meiosis.     

小麦异附加系种质资源的开发利用初探

为了开发利用小麦异附加系种质资源,笔者从利用小麦异附加系与相应的单、缺体杂交选育小麦异代换系;利用小麦异附加系与拟斯卑尔脱山羊草杂交,诱导部分同源染色体配对重组选育异代换系;利用小麦异附加系与Ph隐性突变体Ph1b杂交选育易位系;利用小麦异附加系进行组织培养选育异易位系;利用小麦异附加系进行辐射诱发异源易位;利用小麦异附加系可以选育小麦核不育系的XYZ系方面探讨了利用途径.     

Evidence for reduced recombination on the nondisjoined chromosomes 21 in Down syndrome

Trisomy 21 usually results from nondisjunction during meiosis I. In order to determine whether nondisjunction results from failure of normal chromosome pairing or premature unpairing, recombination frequencies were estimated between DNA polymorphic markers on the long arm of chromosome 21 in families containing one individual with trisomy 21. The recombination frequencies on chromosomes 21 that had undergone nondisjunction were then compared to those on chromosomes 21 that had disjoined normally. The data indicate that recombination is reduced between DNA markers on nondisjoined chromosomes 21. These results are consistent with the hypothesis that reduced chiasma formation predisposes to nondisjunction, resulting in trisomy 21 in humans.     

Cd胁迫诱导拟南芥幼苗DNA损伤分析

以拟南芥为供试植物,通过基于随机引物扩增多态性(RAPD)法的DNA损伤分析,酶联免疫吸附(ELISA)法的DNA甲基化分析以及Real-time PCR的DNA损伤修复与细胞周期相关基因的表达分析,研究了Cd(0、0.125、0.25、1.0、2.5 mg·L~(-1))胁迫5 d的拟南芥幼苗DNA损伤、DNA损伤修复系统以及细胞周期对胁迫的响应。结果显示,随Cd浓度的增加DNA损伤加剧,全基因组甲基化水平较对照组显著增加(P0.01或P0.05),细胞周期调控基因PCNA1、PCNA2,错配修复(MMR)基因MLH1、MSH_2、MSH6,非同源末端连接(NHEJ)标志基因KU70、MRE11、GR1,同源重组(HR)标志基因RAD51、BRCA1的表达均与Cd胁迫浓度呈明显的倒U型剂量效应关系,DNA修复系统对Cd胁迫的敏感性依次为MMRHRNHEJ。该结果表明:轻度Cd胁迫主要引起DNA错配损伤,并且该损伤易修复;随着Cd胁迫的增强,会引起DNA断裂与染色体损伤,损伤较难修复。另外,对Cd胁迫响应最敏感的MSH6、MLH1基因可作为表征Cd胁迫对于拟南芥遗传毒性效应的有效生物标记物。     

基因重组技术在猪伪狂犬病毒基因工程疫苗中的研究进展

猪伪狂犬病是一种由伪狂犬病病毒(Pseudorabies virus,PRV)感染引起的急性高度接触性传染病,可造成不同日龄的猪发病,给养猪业造成了严重的经济损失.预防、控制甚至消灭猪伪狂犬病的主要措施之一是疫苗免疫接种.通过基因重组技术对PRV关键毒力基因进行改造,敲除毒力基因或插入免疫增强基因,是降低PRV毒力或提...     

山西太岳山核桃楸光合特性的研究

用Li 6 4 0 0光合作用测定系统对自然生长的核桃楸壮龄林木的光合作用进行测定 .结果如下 :①核桃楸的光合速率在 1d内有两个峰值 ,两个光合峰高度接近 ,上午的光合速率峰持续时间较下午的长 .叶片气孔导度、蒸腾速率与光合速率间表现出较强的正相关 ,日变化曲线呈双峰型 .叶片胞内CO2 浓度受气孔导度和大气CO2 浓度的双重影响 ,呈现出早晚大幅升高 ,正午有 1h的峰值的日变化进程 .②在控制条件下 ,叶片的光合速率在低光合有效辐射 (0～ 5 0μmol (m2 ·s)下 ,表现出随CO2 浓度增加而降低的趋势 ;高出此光合有效辐射值后 ,光合速率随CO2 浓度的增加呈抛物线变化 ,最高点对应的光合有效辐射下的CO2 饱和点 ,且随光合有效辐射的增加CO2 饱和点增高 .当光合有效辐射为0～ 12 0 0 μmol (m2 ·s)时 ,叶片的气孔导度表现出随光合有效辐射的增加而增大 ,随CO2 浓度的增加而减小的趋势 ;光合有效辐射超过 12 0 0 μmol (m2 ·s)后 ,气孔导度随光合有效辐射、CO2 浓度的增加而降低 .叶片蒸腾速率的变化趋势与气孔导度类似 .     

The Meiotic Behavior of an Alien Chromosome in Triticum aestivum-Haynaldia villosa Monosomic Addition Lines

By the combination of cytological analysis and using genomic in situ hybridization technique to identify an alien chromosome in wheat-Haynaldia villosa monosomic addition lines, we studied the meiotic behavior of the alien chromosome. The results indicated that the frequency of bivalent pairing was lower than the value expected in PMCs of two monosomic addition lines, the frequency of wheat chromosomes unpairing increased, and the wheat homologous chromosome pairing was interfered with by the added chromosome 6V at metaphase I. The chromosome 6V lagged in 20.3% -29.3% of PMCs, sister chromatids 6V early divided in 29.0% - 34.1% of PMCs, the single chromosome 6V in 18.2% - 26.1% of PMCs went to a pole randomly,the breakage frequency of chromosome 6V was 1.2% - 2.9%. Meanwhile, it was also found that several wheat chromosomes showed earlier division, lagging and breakage in a few PMCs. It revealed that the added chromosome 6V influenced the behavior of wheat chromosomes at anaphase. It was also found that the translocation was produced between 6V and wheat chromosomes in 1.2% of PMCs. It offered evidence for translocation between wheat and Haynaldia villosa 6V chromosomes.