MEIOREC_Meiotic Recombination in Plants: controlling the transition of DNA double-strand breaks to genetic crossovers

MEIOREC_植物减数分裂重组：控制DNA双链断裂到遗传交叉的转变

• 批准号: 354617974
• 负责人: Professorin Dr. Mathilde Grelon
• 金额: --
• 依托单位: Research Institute of Molecular Pathology (IMP)
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2018
• 资助国家: 德国
• 起止时间: 2017-12-31 至 2021-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/354617974?language=en
• 关键词: MEIOREC_Meiotic; Recombination; Plants; controlling; transition

Genetic variation is generated through homologous recombination during meiosis and underpins plant breeding efforts to deliver the rapid improvements in crops that will be required to ensure Food Security. HR is initiated by the formation of DNA double-strand breaks (DSBs) by the SPO11 complex. DSBs are processed by components of the HR pathway where they are repaired as crossovers (COs), which recombine the homologous parental chromosomes, or non-crossovers (NCOs), where only short stretches of DNA are exchanged. In plants most DSBs are repaired as NCOs. Moreover, the distribution of COs, notably in cereal crops, is localized to particular chromosomal regions. These limitations significantly lessen the genetic variation that can be generated in each meiotic division. Extensive studies by the MEIOREC investigators have led to significant progress in understanding the basis of these limitations and how they may be addressed. Nevertheless, a full understanding of the factors that control the transition of a DSB at a particular genomic locus to a CO and how this can be optimized has not yet been elucidated. We aim to decipher the molecular steps involved in the DSB to CO transition and evaluate strategies to manipulate CO formation using model species, thereby laying the foundation for subsequent translation of the most promising into crops. Our research will focus on three different stages in the control of CO formation in plants: (i) Factors controlling DSB formation: We will determine their minimum requirements. We will evaluate the potential of a MTOPVIB-CRISPR system as the basis for targeting DSBs complexes to recombination-cold regions. We will investigate the use of a suppression cassette (dCas9) and multiple short guide RNAs of the CO suppressor genes to enhance CO frequency. (ii) DSB processing, stable joint molecule formation and CO resolution: We will address how early steps in DSB processing and recombinase loading are linked and how this impacts on the CO/non-CO decision by analysing the role of the MRN complex and COM1 in the efficient removal of SPO11 from the DSB ends that is essential for loading of the DMC1 and RAD51 recombinases. We will also test the hypothesis that efficiency of CO formation in the recombination reaction can be influenced by Cas9-mediated targeting of the MUS81/GEN1 resolvases. (iii) Relationship of chromosome remodelling and CO formation: We will analyse the influence of posttranslational modifications of the chromosome axis in relation to CO formation and distribution. The inter-relationship between extensive remodelling of the chromosome axis and synaptonemal complex at the leptotene/zygotene transition and the maturation of CO designated recombination intermediates will be also analysed. The link between the programmed remodelling of the chromosome axis and CO distribution will be further investigated in barley and maize.

遗传变异是通过减数分裂过程中的同源重组产生的，并支持植物育种工作，以实现确保粮食安全所需的作物快速改良。HR由SPO 11复合物形成DNA双链断裂（DSB）启动。DSB由HR途径的组分处理，在那里它们被修复为交叉（CO），其重组同源亲本染色体，或非交叉（NCO），其中仅交换短段DNA。在工厂中，大多数DSB作为NCO进行维修。此外，CO的分布，特别是在谷类作物中，定位于特定的染色体区域。这些限制大大减少了在每次减数分裂中可能产生的遗传变异。MEIOREC调查人员的广泛研究在理解这些限制的基础以及如何解决这些限制方面取得了重大进展。然而，控制特定基因组位点处的DSB向CO的转变的因素以及如何优化这一点的充分理解尚未得到阐明。我们的目标是破译参与DSB到CO过渡的分子步骤，并评估使用模型物种操纵CO形成的策略，从而为随后将最有前途的转化为作物奠定基础。我们的研究将集中在三个不同的阶段，在控制CO的形成在植物：（i）控制DSB形成的因素：我们将确定其最低要求。我们将评估MTOPVIB-CRISPR系统作为将DSB复合物靶向重组冷区域的基础的潜力。我们将研究使用抑制盒（dCas 9）和CO抑制基因的多个短向导RNA来提高CO频率。(ii)DSB加工，稳定的联合分子形成和CO分辨率：我们将通过分析MRN复合物和COM 1在从DSB末端有效去除SPO 11中的作用来解决DSB加工和重组酶加载的早期步骤如何联系以及这如何影响CO/非CO决定，这对于加载DMC 1和RAD 51重组酶至关重要。我们还将测试重组反应中CO形成的效率可以受到Cas9介导的MUS 81/GEN 1解离酶靶向的影响的假设。(iii)染色体重塑和CO形成的关系：我们将分析染色体轴的翻译后修饰对CO形成和分布的影响。广泛重塑的染色体轴和联会复合体在细线期/偶线期过渡和CO指定的重组中间体的成熟之间的相互关系也将进行分析。在大麦和玉米中，将进一步研究染色体轴的程序性重塑与CO分布之间的联系。