Meiotic prophase length modulates Tel1-dependent DNA double-strand break interference

During meiosis, genetic recombination is initiated by the formation of many DNA double-strand breaks (DSBs) catalysed by the evolutionarily conserved topoisomerase-like enzyme, Spo11, in preferred genomic sites known as hotspots. DSB formation activates the Tel1/ATM DNA damage responsive (DDR) kinase, locally inhibiting Spo11 activity in adjacent hotspots via a process known as DSB interference. Intriguingly, in S. cerevisiae, over short genomic distances (<15 kb), Spo11 activity displays characteristics of concerted activity or clustering, wherein the frequency of DSB formation in adjacent hotspots is greater than expected by chance. We have proposed that clustering is caused by a limited number of sub-chromosomal domains becoming primed for DSB formation. Here, we provide evidence that DSB clustering is abolished when meiotic prophase timing is extended via deletion of the NDT80 transcription factor. We propose that extension of meiotic prophase enables most cells, and therefore most chromosomal domains within them, to reach an equilibrium state of similar Spo11-DSB potential, reducing the impact that priming has on estimates of coincident DSB formation. Consistent with this view, when Tel1 is absent but Ndt80 is present and thus cells are able to rapidly exit meiotic prophase, genome-wide maps of Spo11-DSB formation are skewed towards pericentromeric regions and regions that load pro-DSB factors early—revealing regions of preferential priming—but this effect is abolished when NDT80 is deleted. Our work highlights how the stochastic nature of Spo11-DSB formation in individual cells within the limited temporal window of meiotic prophase can cause localised DSB clustering—a phenomenon that is exacerbated in tel1∆ cells due to the dual roles that Tel1 has in DSB interference and meiotic prophase checkpoint control.在减数分裂过程中，遗传重组的起始是由进化保守的拓扑异构酶样酶Spo11催化形成大量DNA双链断裂（DNA double-strand breaks, DSBs），且该过程发生在被称为热点的偏好性基因组区域内。DSB的形成会激活Tel1/ATM DNA损伤应答（DDR）激酶，并通过一种被称为DSB干扰的过程，在邻近热点区域局部抑制Spo11的活性。有趣的是，在酿酒酵母（S. cerevisiae）中，当基因组距离较短（<15 kb）时，Spo11的活性表现出协同活性或成簇特征：即相邻热点区域的DSB形成频率高于随机预期的水平。我们曾提出，成簇现象是由有限数量的亚染色体结构域被预先激活以进行DSB形成所导致的。本研究提供证据表明，当通过敲除NDT80转录因子延长减数分裂前期时长时，DSB成簇现象会被消除。我们推测，减数分裂前期的延长可使绝大多数细胞及其内部的大多数染色体结构域达到具有相似Spo11-DSB形成潜能的平衡状态，从而削弱预先激活对共现DSB形成频率估算的影响。与这一观点一致的是，当Tel1缺失但Ndt80存在时，细胞能够快速退出减数分裂前期，此时Spo11-DSB形成的全基因组图谱会偏向着丝粒附近区域以及早期加载促DSB因子的区域——这些区域正是优先发生预先激活的区域——但当NDT80被敲除后，这一效应会消失。本研究揭示了，在减数分裂前期的有限时间窗口内，单个细胞中Spo11介导的DSB形成的随机性如何导致局部DSB成簇现象；而在tel1Δ细胞中，由于Tel1同时参与DSB干扰和减数分裂前期检查点调控，这一现象会被进一步加剧。