Meiotic chromosome synapsis and recombination in yeast.

描述（由申请人提供）：本申请涉及酵母减数分裂期间同源染色体之间的相互作用。(I)DSB独立配对。我们将使用FISH来探测DSB独立的同源配对相对丰富的R-和G-带，姐妹染色单体凝聚力和染色质结构蛋白的参与的关系。我们将使用“捕获染色体构象”（3C）方法来识别配对接触的位点。我们将进行一个试点实验，以研究一种可能的新的配对缺陷突变体的测定。(II)重组的开始：DSB转变。我们将通过3C方法学和遗传学研究来调查前DSB重组体是否与其潜在的染色体轴物理相关。我们还将使用3C方法来识别新生的DSB/合作伙伴的相互作用，并探讨同源/姐妹歧视。我们将进一步探索控制任何给定减数分裂核中单个位点可能发生的DSB数量的限制。(III)重组的后期阶段。我们将进一步探讨影响交换控制过渡的突变体中的减数分裂，并注意温度的重要影响。我们将继续分析野生型和选择的突变体的花束阶段。我们将继续分析Mlh 3在减数分裂重组中的作用。我们将研究突变的表型，怀疑影响转换的单端入侵双霍利迪路口在粗线期中期。我们将继续研究哪些拓扑异构体的双霍利迪连接发生在减数分裂。(IV)减数分裂染色体结构和力学。我们将进一步探讨我们最近的研究揭示的染色质/轴/姐妹的相互作用。我们将使用3C方法来研究中前期染色体的物理性质。我们将在体内研究联会复合体扭转的动力学。我们将开始发展在体外分离、分析和物理操作粗线期染色体的方法。

DESCRIPTION (provided by applicant): This application addresses the interactions between homologous chromosomes during meiosis in yeast. (I) DSB-independent pairing. We will use FISH to probe DSB-independent homolog pairing with respect to relative abundance in R- and G-bands, relationship to sister chromatid cohesion and involvement of chromatin structure proteins. We will use "Capturing Chromosome Conformation" (3C) methodology to identify sites of pairing contacts. We will carry out a pilot experiment to investigate a possible new assay for pairing-defective mutants. (II) Initiation of recombination: the DSB transition. We will investigate whether pre-DSB recombinosomes are physically associated with their underlying chromosome axes by 3C methodology and genetic studies. We will also use 3C methodology to identify nascent DSB/partner interactions and to explore homolog/sister discrimination. We will further explore constraints governing the number of DSBs that can occur at a single locus in any give meiotic nucleus. (III) Later stages of recombination. We will further explore meiosis in mutants that affect the crossover control transition, with attention to important effects of temperature. We will continue analysis of the bouquet stage in wild type and selected mutants. We will continue analysis of the role of Mlh3 for meiotic recombination. We will examine the phenotypes of mutations suspected to affect conversion of single-end invasions to double Holliday junctions at mid-pachytene. And we will continue to investigate which topological isomer(s) of double Holliday junctions occur during meiosis. (IV) Meiotic chromosome structure and mechanics. We will further explore chromatin/axis/sister interplay revealed by our recent studies. We will use 3C methodology to investigate physical properties of mid-prophase chromosomes. We will examine the dynamics of synaptonemal complex twisting in vivo. We will begin to develop methods for isolating, analyzing and physically manipulating pachytene chromosomes in vitro.