Single-cell long-read Hi-C, scNanoHi-C2, details 3D genome reorganization in embryonic-stage germ cells.

During mouse development, embryonic-stage germ cells (EGCs) make crucial fate decisions, with female EGCs embarking on meiosis whereas male EGCs enter mitotic arrest until birth. Despite increasing understanding of the reprogramming of epigenetic modifications, the dynamics of three-dimensional (3D) genome structures within individual EGCs remains elusive. Here we present a single-cell input, long-read Hi-C method, termed scNanoHi-C2. We use scNanoHi-C2 to systematically dissect the dynamics of EGC chromatin structures. We find that, despite changes in autosomes similar to spermatogenesis, the X chromosomes of female EGCs show enhanced specific interactions between B compartments. By reconstructing 3D genome models, we observe dynamic chromosome positioning during meiosis, showing that the neighborhood between nonhomologous chromosomes of EGCs is relatively random. Simultaneously, transposable elements undergo dramatic chromatin reorganization and display an asymmetric distribution of Alu/B2 elements around meiotic topologically associated domain boundaries. Moreover, we find that high-order interactions in EGCs at the mitosis stage are mainly enriched in the B compartment, whereas, after the mitosis-to-meiosis transition, enriched high-order interactions shift to refined A compartments, to potentially promote meiotic-specific transcription programs during global genomic condensation. We also reveal an unexpected chromatin structure in mitotic-arrested male EGCs distinct from the previously assumed G0 status, which may prime the unique genome structure for subsequent spermatogenesis. Altogether, our study highlights the potential of scNanoHi-C2 and reveals key features of the chromatin structure reprogramming in EGCs.