Nucleome programming is required for the foundation of totipotency in mammalian germline development [Hi-C]. Nucleome programming is required for the foundation of totipotency in mammalian germline development [Hi-C]

Germ cells are unique in engendering totipotency, yet the mechanisms underlying this capacity remain elusive. Here, we perform comprehensive and in-depth nucleome analysis of mouse germ-cell development in vitro, encompassing pluripotent precursors, primordial germ cells (PGCs) before and after epigenetic reprogramming, and spermatogonia/spermatogonial stem cells (SSCs). Although epigenetic reprogramming, including genome-wide DNA de-methylation, creates broadly open chromatin with abundant enhancer-like signatures, the augmented chromatin insulation safeguards transcriptional fidelity. These insulatory constraints are then erased en masse for spermatogonial development. Notably, despite distinguishing epigenetic programming, including global DNA re-methylation, the PGCs-to-spermatogonia/SSCs development entails further euchromatization. This accompanies substantial erasure of lamina-associated domains, generating spermatogonia/SSCs with a minimal peripheral attachment of chromatin except for pericentromeres—an architecture conserved in primates. Accordingly, faulty nucleome maturation, including persistent insulation and improper euchromatization, leads to impaired spermatogenic potential. Given that PGCs after epigenetic reprogramming serve as oogenic progenitors as well, our findings elucidate a principle for the nucleome programming that creates gametogenic progenitors in both sexes, defining a basis for nuclear totipotency. Overall design: Nucleome profiling of various cell types along murine germ-cell development using Hi-C

生殖细胞在催生细胞全能性方面独具特性，但其具备该能力的分子机制仍未完全阐明。本研究对体外培养的小鼠生殖细胞发育进程开展了全面且深入的核组（nucleome）分析，涵盖多能前体细胞、表观重编程前后的原始生殖细胞（primordial germ cells, PGCs）以及精原细胞/精原干细胞（spermatogonial stem cells, SSCs）。尽管表观重编程（包括全基因组DNA去甲基化）可形成广泛开放的染色质并伴随大量增强子样特征，但增强的染色质绝缘作用可保障转录保真度。而在精原细胞发育阶段，这类绝缘约束会被大规模清除。值得注意的是，尽管经历了包括全基因组DNA重新甲基化在内的特异性表观编程，从PGCs向精原细胞/SSCs的发育过程仍伴随进一步的常染色质化进程。该过程伴随核纤层结合结构域（lamina-associated domains）的大规模清除，使得精原细胞/SSCs的染色质仅在着丝粒周边区域存在少量外周附着，这一染色质架构在灵长类中具有保守性。因此，核组成熟异常（包括绝缘作用持续存在以及常染色质化异常）会导致生精潜能受损。鉴于表观重编程后的PGCs同时可作为卵原前体细胞，本研究的发现阐明了一套核组编程的通用原则：该原则可在两性中生成配子发生前体细胞，为细胞核全能性奠定了理论基础。实验设计概述：通过Hi-C技术对小鼠生殖细胞发育进程中的多种细胞类型开展核组谱型分析。