MCM8 Is Required for a Pathway of Meiotic Double-Strand Break Repair Independent of DMC1 in <em>Arabidopsis thaliana</em>

Mini-chromosome maintenance (MCM) 2–9 proteins are related helicases. The first six, MCM2–7, are essential for DNA replication in all eukaryotes. In contrast, MCM8 is not always conserved in eukaryotes but is present in Arabidopsis thaliana. MCM8 is required for 95% of meiotic crossovers (COs) in Drosophila and is essential for meiosis completion in mouse, prompting us to study this gene in Arabidopsis meiosis. Three allelic Atmcm8 mutants showed a limited level of chromosome fragmentation at meiosis. This defect was dependent on programmed meiotic double-strand break (DSB) formation, revealing a role for AtMCM8 in meiotic DSB repair. In contrast, CO formation was not affected, as shown both genetically and cytologically. The Atmcm8 DSB repair defect was greatly amplified in the absence of the DMC1 recombinase or in mutants affected in DMC1 dynamics (sds, asy1). The Atmcm8 fragmentation defect was also amplified in plants heterozygous for a mutation in either recombinase, DMC1 or RAD51. Finally, in the context of absence of homologous chromosomes (i.e. haploid), mutation of AtMCM8 also provoked a low level of chromosome fragmentation. This fragmentation was amplified by the absence of DMC1 showing that both MCM8 and DMC1 can promote repair on the sister chromatid in Arabidopsis haploids. Altogether, this establishes a role for AtMCM8 in meiotic DSB repair, in parallel to DMC1. We propose that MCM8 is involved with RAD51 in a backup pathway that repairs meiotic DSB without giving CO when the major pathway, which relies on DMC1, fails.微染色体维持蛋白（Mini-chromosome maintenance, MCM）2~9 均为相关解旋酶（helicase）。其中前六个亚型MCM2~7是所有真核生物（eukaryote）DNA复制过程的必需因子。与之不同的是，MCM8并非在所有真核生物中都保守，但拟南芥（Arabidopsis thaliana）中存在该蛋白。在果蝇中，MCM8是95%减数分裂交叉（meiotic crossovers, COs）形成所必需的基因，且对小鼠减数分裂的完成至关重要，这促使我们在拟南芥中开展该基因的减数分裂功能研究。 三个等位基因Atmcm8突变体在减数分裂过程中会出现轻度染色体碎片化表型。该缺陷的产生依赖于程序性减数分裂双链断裂（programmed meiotic double-strand break, DSB）的形成，提示AtMCM8参与减数分裂DSB的修复过程。与之相反，遗传学与细胞学实验均证实，减数分裂交叉的形成并未受到影响。 Atmcm8的DSB修复缺陷在缺失DMC1重组酶（recombinase）或DMC1动态调控异常的突变体（sds、asy1）中会显著加剧。当重组酶DMC1或RAD51其中一个基因发生杂合突变时，Atmcm8突变体的染色体碎片化缺陷同样会被放大。 最后，在无同源染色体的单倍体背景下，AtMCM8的突变也会引发轻度染色体碎片化。而DMC1的缺失会进一步加剧该碎片化程度，这表明在拟南芥单倍体中，MCM8和DMC1均可介导基于姐妹染色单体的DNA修复。 综上，本研究明确了AtMCM8在减数分裂DSB修复中的功能，其作用与DMC1相互平行。我们提出，当依赖DMC1的主要修复通路失效时，MCM8可与RAD51共同参与一条备用修复通路，完成减数分裂DSB的修复且不产生减数分裂交叉。