SPOCD1 is a novel executor of piRNA-directed DNA methylation that links MIWI2 to the de novo DNA methylation machinery. SPOCD1 is a novel executor of piRNA-directed DNA methylation that links MIWI2 to the de novo DNA methylation machinery

In mammals, the acquisition of the germline from the soma provides the germline with an essential challenge, the necessity to erase and reset genomic methylation. De novo genome methylation re-encodes the epigenome including transposable element (TE) silencing. In the male germline RNA-directed DNA methylation silences young active TEs. The PIWI protein MIWI2 (PIWIL4) and its associated PIWI-interacting RNA (piRNAs) act to tether MIWI2 to nascent TE transcripts and instruct DNA methylation of the active TE. The mechanism by which MIWI2 directs de novo TE methylation is poorly understood but central to the immortality of the germline. Here, we define the interactome of MIWI2 in foetal gonocytes that are undergoing de novo genome methylation and identify a novel MIWI2-associated factor SPOCD1 that is essential for TE silencing. The loss of Spocd1 in mice phenocopies that of Miwi2-deficient mice and does not impact on piRNA biogenesis nor localization of MIWI2 to the nucleus. SPOCD1 is a nuclear protein and its expression is restricted to the period of de novo genome methylation. We found SPOCD1 co-purified in vivo with constituents of several repressive chromatin remodelling complexes (NURD and BAF) as well as DNMT3L and DNMT3A, components of the de novo methylation machinery. We propose a model whereby tethering of MIWI2 to a nascent TE transcript recruits repressive chromatin remodelling activities and the de novo methylation apparatus through its association with SPOCD1. In summary, we have identified a novel and essential executor of mammalian piRNA-directed DNA methylation.

在哺乳动物中，生殖系从体细胞中获取的过程对生殖系构成一项核心挑战——必须清除并重置基因组甲基化。从头基因组甲基化会对表观基因组进行重编码，其中涵盖转座因子（transposable element, TE）的沉默。在雄性生殖系中，RNA指导的DNA甲基化可沉默年轻的活跃转座因子。PIWI蛋白MIWI2（PIWIL4）及其结合的PIWI互作RNA（PIWI-interacting RNA, piRNAs）可将MIWI2锚定在新生TE转录本上，并指导活跃转座因子的DNA甲基化。MIWI2介导从头转座因子甲基化的具体机制目前仍不甚明晰，却是维持生殖系不朽性的核心环节。本研究针对正在经历从头基因组甲基化的胎儿生殖母细胞，解析了MIWI2的互作组，并鉴定出一种全新的、对转座因子沉默至关重要的MIWI2结合因子SPOCD1。小鼠中Spocd1基因的缺失表型与Miwi2缺陷小鼠完全一致，且既不影响piRNA的生成，也不会干扰MIWI2向细胞核的定位。SPOCD1是一种核蛋白，其表达仅局限于从头基因组甲基化发生的时段。实验发现，SPOCD1在体内可与多种抑制性染色质重塑复合物（NURD与BAF）的组分，以及从头甲基化装置的核心组分DNMT3L和DNMT3A共纯化。据此我们提出模型：MIWI2与新生转座因子转录本结合后，可通过与SPOCD1的相互作用，招募抑制性染色质重塑活性与从头甲基化装置。综上，本研究鉴定出了哺乳动物piRNA指导的DNA甲基化过程中一种全新且不可或缺的执行因子。