Data from: Mek1 down regulates Rad51 activity during yeast meiosis by phosphorylation of Hed1

During meiosis, programmed double strand breaks (DSBs) are repaired preferentially between homologs to generate crossovers that promote proper chromosome segregation at Meiosis I. In many organisms, there are two strand exchange proteins, Rad51 and the meiosis-specific Dmc1, required for interhomolog (IH) bias. This bias requires the presence, but not the strand exchange activity of Rad51, while Dmc1 is responsible for the bulk of meiotic recombination. How these activities are regulated is less well established. In dmc1Δ mutants, Rad51 is actively inhibited, thereby resulting in prophase arrest due to unrepaired DSBs triggering the meiotic recombination checkpoint. This inhibition is dependent upon the meiosis-specific kinase Mek1 and occurs through two different mechanisms that prevent complex formation with the Rad51 accessory factor Rad54: (i) phosphorylation of Rad54 by Mek1 and (ii) binding of Rad51 by the meiosis-specific protein Hed1. An open question has been why inhibition of Mek1 affects Hed1 repression of Rad51. This work shows that Hed1 is a direct substrate of Mek1. Phosphorylation of Hed1 at threonine 40 helps suppress Rad51 activity in dmc1Δ mutants by promoting Hed1 protein stability. Rad51-mediated recombination occurring in the absence of Hed1 phosphorylation results in a significant increase in non-exchange chromosomes despite wild-type levels of crossovers, confirming previous results indicating a defect in crossover assurance. We propose that Rad51 function in meiosis is regulated in part by the coordinated phosphorylation of Rad54 and Hed1 by Mek1.

在减数分裂过程中，程序性双链断裂（double strand breaks, DSBs）优先在同源染色体之间完成修复，以产生交叉互换，保障减数第一次分裂时染色体的正常分离。在多数生物体内，存在两种链交换蛋白：Rad51与减数分裂特异性蛋白Dmc1，二者均为同源染色体间重组偏向性（interhomolog, IH）所必需。该偏向性依赖于Rad51的存在，却无需其链交换活性；而Dmc1则负责绝大多数减数分裂重组事件。目前，这些蛋白活性的调控机制尚未被充分阐明。在dmc1Δ突变体中，Rad51会被主动抑制，未修复的DSBs会激活减数分裂重组检验点，最终导致细胞发生前期阻滞。这种抑制作用依赖于减数分裂特异性激酶Mek1，并通过两种不同的途径阻止Rad51与其辅助因子Rad54形成复合物：(i) Mek1对Rad54的磷酸化修饰；(ii) 减数分裂特异性蛋白Hed1与Rad51结合。长期以来，一个悬而未决的科学问题是，为何抑制Mek1的活性会影响Hed1对Rad51的抑制作用。本研究证实，Hed1是Mek1的直接底物。Hed1在苏氨酸40位点的磷酸化，可通过提升Hed1的蛋白质稳定性，在dmc1Δ突变体中抑制Rad51的活性。当缺失Hed1的磷酸化修饰时，Rad51介导的重组过程虽能维持野生型水平的交叉互换，却会导致非交换染色体数量显著增加，这一结果验证了此前关于交换保证存在缺陷的研究结论。我们提出，减数分裂中Rad51的功能，部分通过Mek1对Rad54与Hed1的协同磷酸化得以调控。