Garvan Institute of Medical Research, Darlinghurst, Sydney, NSW, 2010, Australia.
St Vincent's Clinical School, Faculty of Medicine, University of New South Wales, Kensington, Sydney, NSW, 2010, Australia.
EMBO J. 2024 Oct;43(20):4786-4804. doi: 10.1038/s44318-024-00210-5. Epub 2024 Aug 29.
DNA i-motif structures are formed in the nuclei of human cells and are believed to provide critical genomic regulation. While the existence, abundance, and distribution of i-motif structures in human cells has been demonstrated and studied by immunofluorescent staining, and more recently NMR and CUT&Tag, the abundance and distribution of such structures in human genomic DNA have remained unclear. Here we utilise high-affinity i-motif immunoprecipitation followed by sequencing to map i-motifs in the purified genomic DNA of human MCF7, U2OS and HEK293T cells. Validated by biolayer interferometry and circular dichroism spectroscopy, our approach aimed to identify DNA sequences capable of i-motif formation on a genome-wide scale, revealing that such sequences are widely distributed throughout the human genome and are common in genes upregulated in G0/G1 cell cycle phases. Our findings provide experimental evidence for the widespread formation of i-motif structures in human genomic DNA and a foundational resource for future studies of their genomic, structural, and molecular roles.
DNA i-motif 结构存在于人类细胞的核内，被认为对基因组调控起着关键作用。虽然免疫荧光染色，以及最近的 NMR 和 CUT&Tag 已经证明和研究了 i-motif 结构在人类细胞中的存在、丰度和分布，但这些结构在人类基因组 DNA 中的丰度和分布仍不清楚。在这里，我们利用高亲和力 i-motif 免疫沉淀结合测序的方法，对 MCF7、U2OS 和 HEK293T 细胞的纯化基因组 DNA 中的 i-motif 进行了定位。通过生物层干涉测量法和圆二色光谱法验证，我们的方法旨在鉴定能够在全基因组范围内形成 i-motif 的 DNA 序列，结果表明这些序列广泛分布于人类基因组中，并且在 G0/G1 细胞周期阶段上调的基因中很常见。我们的研究结果为 i-motif 结构在人类基因组 DNA 中的广泛形成提供了实验证据，并为它们在基因组、结构和分子水平上的作用的进一步研究提供了基础资源。
