Multiple Pairwise Analysis of Non-homologous Centromere Coupling Reveals Preferential Chromosome Size-Dependent Interactions and a Role for Bouquet Formation in Establishing the Interaction Pattern

During meiosis, chromosomes undergo a homology search in order to locate their homolog to form stable pairs and exchange genetic material. Early in prophase, chromosomes associate in mostly non-homologous pairs, tethered only at their centromeres. This phenomenon, conserved through higher eukaryotes, is termed centromere coupling in budding yeast. Both initiation of recombination and the presence of homologs are dispensable for centromere coupling (occurring in spo11 mutants and haploids induced to undergo meiosis) but the presence of the synaptonemal complex (SC) protein Zip1 is required. The nature and mechanism of coupling have yet to be elucidated. Here we present the first pairwise analysis of centromere coupling in an effort to uncover underlying rules that may exist within these non-homologous interactions. We designed a novel chromosome conformation capture (3C)-based assay to detect all possible interactions between non-homologous yeast centromeres during early meiosis. Using this variant of 3C-qPCR, we found a size-dependent interaction pattern, in which chromosomes assort preferentially with chromosomes of similar sizes, in haploid and diploid spo11 cells, but not in a coupling-defective mutant (spo11 zip1 haploid and diploid yeast). This pattern is also observed in wild-type diploids early in meiosis but disappears as meiosis progresses and homologous chromosomes pair. We found no evidence to support the notion that ancestral centromere homology plays a role in pattern establishment in S. cerevisiae post-genome duplication. Moreover, we found a role for the meiotic bouquet in establishing the size dependence of centromere coupling, as abolishing bouquet (using the bouquet-defective spo11 ndj1 mutant) reduces it. Coupling in spo11 ndj1 rather follows telomere clustering preferences. We propose that a chromosome size preference for centromere coupling helps establish efficient homolog recognition.

在减数分裂过程中，染色体需进行同源搜索以定位其同源染色体，形成稳定配对并交换遗传物质。减数分裂前期早期，染色体多以非同源配对形式结合，且仅通过着丝粒相互拴系。这一在高等真核生物中保守存在的现象，在出芽酵母中被称为着丝粒偶联。着丝粒偶联并不依赖重组起始与同源染色体的存在（该过程可发生于spo11突变体以及经诱导进入减数分裂的单倍体中），但必须依赖联会复合体（synaptonemal complex, SC）蛋白Zip1的存在。着丝粒偶联的本质与机制仍有待阐明。本研究首次针对着丝粒偶联开展成对分析，以期揭示这些非同源相互作用中可能存在的内在规律。我们设计了一种基于染色体构象捕获（chromosome conformation capture, 3C）的新型检测方法，用于检测减数分裂早期酵母非同源着丝粒之间的所有潜在相互作用。利用这一改良的3C-qPCR技术，我们在单倍体与二倍体spo11细胞中发现了一种尺寸依赖性的相互作用模式：染色体优先与尺寸相近的染色体配对，但在着丝粒偶联缺陷的突变体（spo11 zip1单倍体与二倍体酵母）中未观察到此现象。该模式在减数分裂早期的野生型二倍体酵母中同样存在，但随着减数分裂进程推进与同源染色体配对完成，该模式会逐渐消失。我们未发现任何证据支持‘酿酒酵母（S. cerevisiae）全基因组复制后，祖先着丝粒同源性在该相互作用模式的建立中发挥作用’这一观点。此外，我们发现减数分裂花束结构在着丝粒偶联的尺寸依赖性建立中发挥作用：破坏花束结构（通过使用花束缺陷型spo11 ndj1突变体）会削弱该尺寸依赖性。而spo11 ndj1突变体中的着丝粒偶联转而遵循端粒簇集偏好。我们提出，着丝粒偶联的染色体尺寸偏好性有助于实现高效的同源染色体识别。