DNA cytosine methylation suppresses meiotic recombination at the sex-determining region (H3K27ac ChIP-seq)

In sexual organisms, random meiotic recombination between homologous chromosomes is vital to maximize genetic variation among offspring. The sex-determining region (SDR), however, does not undergo recombination, as required for the maintenance of distinct alleles. The contribution of epigenetic versus genetic mechanisms to suppress recombination in SDRs and how the suppression mechanisms evolve remain poorly understood. Here we describe the mechanistic control of meiotic recombination at the mating-type locus (MT) in the green alga Chlamydomonas reinhardtii. We identify a maintenance DNA methyltransferase, DNMT1, mutation of which leads to the depletion of 95% of 5-methylcytosines (5mCs) in the nuclear genome. While the 5mC deficiency causes no discernible alteration in haploid vegetative growth or sexual differentiation, the dnmt1 homozygotes display substantially reduced spore viability and 4-progeny-tetrad frequency. Strikingly, in dnmt1 homozygotes, anomalous meiotic recombination takes place at MT, generating haploid progenies with mixed mating-type harboring both plus and minus markers. Although the repressive histone methylation H3K9me1 at MT is lost concurrently in dnmt1 strains, loss of histone methylation alone by gene deletion of the responsible histone methyltransferase SET3p does not lead to anomalous recombination at MT. Thus, DNA methylation, rather than histone modification, mediates the recombination suppression at MT in C. reinhardtii. This finding suggests that in early eukaryotes, which likely lacked histone-based chromatin modification, DNA methylation may have been co-opted to suppress meiotic recombination between alleles responsible for separate sexes. Overall design: Chromatin immunoprecipitation sequencing (ChIP-seq) for histone modifications H3K27ac in WT, dnmt1 and set3 strains.

在有性生殖生物中，同源染色体间的随机减数分裂重组对最大化子代的遗传变异至关重要。然而，性别决定区域（sex-determining region, SDR）并不会发生重组，这是维持差异化等位基因所必需的调控机制。目前，表观遗传与遗传机制对SDR内重组抑制的贡献，以及此类抑制机制的演化路径，仍未被充分阐明。本研究针对莱茵衣藻（Chlamydomonas reinhardtii）的交配型位点（mating-type locus, MT），阐明了其减数分裂重组的调控机制。本研究鉴定出一种维持型DNA甲基转移酶（DNA methyltransferase, DNMT1），其突变可导致核基因组中95%的5-甲基胞嘧啶（5-methylcytosine, 5mC）被清除。尽管5mC缺失不会对单倍体营养生长或有性分化产生可检测到的影响，但dnmt1纯合子的孢子存活率与4子代四分体频率均显著降低。令人意外的是，在dnmt1纯合子中，MT位点处出现了异常的减数分裂重组，产生了同时携带正、负标记的混合交配型单倍性子代。尽管dnmt1突变株中MT位点的抑制性组蛋白甲基化修饰H3K9me1同时丧失，但仅通过敲除负责的组蛋白甲基转移酶SET3p来去除组蛋白甲基化，并不会导致MT位点出现异常重组。因此，在莱茵衣藻中，是DNA甲基化而非组蛋白修饰介导了MT位点的重组抑制。该发现提示，在早期真核生物——这类生物大概率缺乏基于组蛋白的染色质修饰——中，DNA甲基化可能被征用来抑制负责雌雄分化的等位基因之间的减数分裂重组。实验整体设计：对野生型（wild type, WT）、dnmt1及set3菌株的组蛋白修饰H3K27ac进行染色质免疫沉淀测序（chromatin immunoprecipitation sequencing, ChIP-seq）。