Super-sized clustered loops anchored at transposon Helitrons in Xenopus tropicalis sperm associate with late gene expression during embryogenesis [ChIP-Seq2]. Super-sized clustered loops anchored at transposon Helitrons in Xenopus tropicalis sperm associate with late gene expression during embryogenesis [ChIP-Seq2]

Vertebrate sperm genome differs from somatic cells and undergoes dramatic transformation after fertilization. However, the functional implications of the sperm genome structures have not been fully investigated. Here we show, in the sperms of Xenopus tropicalis, tens of genomic regions harbor multi-megabases, super-sized clustered loops (SSCLs) whose anchors are enriched with Helitrons, the only group of rolling-circle transposons. SSCL anchors are inaccessible and absent of active histone modifications, implying that SSCLs are repressive in nature. Moreover, genes associated with SSCL anchors express late during development, suggesting 3D structure in sperm may associate with gene expression control during embryo development. The absence of CTCF and RNAPII at SSCL anchors argues against CTCF-mediated or transcription-related looping for the SSCLs establishment. Furthermore, our molecular simulation excludes looping and supports a phase separation model through which SSCLs may form. Taken together, our work reveals a previously undiscovered, repressive 3D structure in sperm that may mediate intergenerational gene regulation. Overall design: Xenopus tropicalis sperm and Xenopus tropiclis x Xenopus laevis hybrid staged embryos were obtained. Hi-C, ChIP-seq, ATAC-seq and RNA-seq were performed in these cells.

脊椎动物精子基因组与体细胞基因组存在显著差异，且在受精后会经历剧烈的染色质重塑过程。然而，目前对于精子基因组三维结构的功能意义尚未得到充分研究。本研究以热带爪蟾（Xenopus tropicalis）精子为研究对象，发现数十个长达数兆碱基的基因组区域携带有超大型簇状环（super-sized clustered loops, SSCLs），其锚定区域富集有赫里逊转座子（Helitrons）——目前已知唯一的滚环转座子类群。SSCL锚定区域无法被染色质开放检测技术捕获，且不存在活跃的组蛋白修饰，这表明SSCLs本质上具有转录抑制功能。此外，与SSCL锚定区域相关的基因多在发育后期表达，这提示精子中的三维基因组结构可能与胚胎发育过程中的基因表达调控存在关联。SSCL锚定区域不存在CCCTC结合因子（CTCF）和RNA聚合酶II（RNAPII）的结合，这表明SSCLs的形成并非由CTCF介导或依赖转录的环化过程所驱动。此外，我们的分子模拟实验排除了传统环化形成的可能，支持SSCLs通过相分离机制形成的模型。综上，本研究揭示了精子中此前未被发现的具有转录抑制功能的三维基因组结构，该结构可能参与跨代基因调控。整体实验设计：我们获取了热带爪蟾（Xenopus tropicalis）精子以及热带爪蟾与非洲爪蟾（Xenopus laevis）杂交的分期胚胎，随后对这些样本进行了Hi-C、染色质免疫共沉淀测序（ChIP-seq）、转座酶可及性测序（ATAC-seq）以及RNA测序（RNA-seq）实验。