Insights from single-molecule force spectroscopy into chromatin topology

Abstract The organization of DNA into chromatin within the eukaryotic nucleus represents one of the most intricate forms of biological packaging. This organization is inherently hierarchical, spanning multiple length scales: from DNA wrapping around histone octamers to form nucleosomes, to the folding of nucleosome arrays and the establishment of higher-order chromatin domains. Understanding how these scales interconnect is essential for explaining how cells regulate gene expression and replicate their genomes. Single-molecule force spectroscopy has provided powerful mechanistic insights into this architecture by enabling direct measurements of the forces that govern chromatin folding and unfolding. These approaches have clarified how chromatin fibers behave under tension and how nucleosome-level interactions contribute to fiber compaction and stability. In this review, we summarize key discoveries enabled by such techniques and discuss emerging opportunities for probing chromatin in increasingly complex and biologically relevant contexts.