DNA双链断裂-融合介导的同源染色体重组分子机理研究

Meiosis is a highly conserved process in the life cycles of all sexually reproductive organisms. It includes one round of DNA replication and two successive rounds of nuclear division. During prophase I of meiosis, recombination between homologous chromosomes occurs, these are then separated at meiosis I, and the two sister chromatids are pulled to the opposite poles during meiosis II. Homologous recombination is essential for the genetic diversity of meiotic products and is also a prerequisite for the correct segregation of homologous chromosomes. Those haploid reproductive cells then develop into gametophytes. After fertilization, the chromosome number of zygotes returns to that of their parents. Therefore, meiosis is important for sexually reproducing organisms. It facilitates the maintenance of the stable chromosome number of a species and more importantly produces genetic variation in gametes, which in turn promote genetic and phenotypic variation in its offspring. ..In most organisms, a stable connection is established by crossovers (COs). COs not only ensure the proper homologous segregation, but create new combinations of alleles leading to increased variation. The number and distribution of COs are tightly controlled. If multiple COs occur along a homologous pair, they are more evenly spaced than would be expected by chance. This phenomenon is known as interference. Although more than 50 genes related to CO formation have identified, the molecular mechanisms on homologous chromosome pairing and recombination are still poorly understood.. .In this proposal, we are going to identify the mutants with defects of either double-strand-break (DSB) initiation or DSB repairing from our rice mutant library, which contains over 300 independent lines all with meiotic defects. By using of those mutants, five genes related to DSB initiation or repairing would be isolated. Their biological functions on homologous recombination would also be carefully investigated. Together with the knowledge on the meiotic genes in rice isolated so far in our lab, those data will strength our understanding of the molecular mechanism on homologous recombination induced by DSB formation and its successful repairing.

减数分裂过程中，同源染色体配对与重组是非常复杂而又关键的生物学过程，它不仅促进来自父母双方的遗传物质充分交流和重组，还使同源染色体间形成交叉结，保证同源染色体在子细胞中的正确分配和分离。因此充分认识这一复杂生物学过程，了解它们的作用网络与调控途径，有助于最终实现对它的遗传调控。本课题将以水稻为模式生物，对本实验室构建的涵盖300多份独立来源水稻减数分裂突变体进行详细细胞学研究，依据减数分裂染色体的配对特征、染色体桥及碎片的有无，以及γH2AX抗体免疫荧光染色反应等手段，筛选和鉴定具有DNA 双链断裂-融合（Double-Strain-Break，DSB）形成或修复缺陷的突变体，进一步分离参与水稻DSB形成与修复的关键基因5个，结合本实验室已克隆的10多个水稻减数分裂相关基因，深入研究由DSB形成与修复介导的Crossover形成机制，为深入探明同源染色体重组分子机制带来新的思路和启示。

减数分裂过程中，同源染色体配对与重组是相当复杂又关键的生物学过程。它不仅促进了来自父母双方的遗传物质能够充分交流和重组，还促使同源染色体间形成交叉结，从而保证同源染色体在子细胞中的正确分配和分离。因此，充分认识这一复杂生物学过程，克隆参与这一过程的关键基因，了解它们的作用网络与调控途径，有助于最终实现对这一复杂生命过程进行遗传操作和调控。本课题重点以水稻为模式生物，克隆鉴定参与DSB形成与同源重组的关键基因15个。表型分析结合基因功能解析，阐明这些基因在减数分裂过程中的详细生物学功能：OsMTOPVIB是一个全新的DSB形成起始因子，参与水稻DSB形成；MRE11、XRCC3、OsHUS1、OsRAD1、OsRAD51C、OsDMC1及新蛋白MEICA1参与水稻DSB早期修复，保障水稻减数分裂同源重组过程；OsMSH4及OsMSH5参与调控水稻同源重组后期交换的发生及交叉结的形成；CRC1及P31comet是水稻联会复合体新组分，在同源配对及联会中起着至关重要的作用；ZYGO1作为新的F-box蛋白，首次被发现调控减数分裂偶线期染色体形态建成； BRK1定位于水稻同源染色体着丝粒处，保证减数第一次同源染色体的正确分离。. 上述相关研究分别从同源重组的起始、修复及完成三个阶段充分解析了单子叶模式植物水稻的减数分裂过程，初步构建由DSB形成与修复介导的同源染色体重组基因网络，完善酵母及动物中减数分裂同源重组模型。同时，对同源重组交换发生及重组频率调节的基础研究亦为育种过程中如何切实提高重组频率奠定了理论基础。本项目执行期间，发表国际主流学术刊物论文17篇，申请相应专利5项，获得授权专利1项。