High-throughput AFM analysis reveals unwrapping pathways of H3 and CENP-A nucleosomes

Konrad, Sebastian F.; Vanderlinden, Willem; Frederickx, Wout; Brouns, Tine; Menze, Björn H.; De Feyter, Steven und Lipfert, Jan (2021): High-throughput AFM analysis reveals unwrapping pathways of H3 and CENP-A nucleosomes. In: Nanoscale, Bd. 13, Nr. 10: S. 5435-5447

Abstract

Nucleosomes, the fundamental units of chromatin, regulate readout and expression of eukaryotic genomes. Single-molecule experiments have revealed force-induced nucleosome accessibility, but a high-resolution unwrapping landscape in the absence of external forces is currently lacking. Here, we introduce a high-throughput pipeline for the analysis of nucleosome conformations based on atomic force microscopy and automated, multi-parameter image analysis. Our data set of similar to 10 000 nucleosomes reveals multiple unwrapping states corresponding to steps of 5 bp DNA. For canonical H3 nucleosomes, we observe that dissociation from one side impedes unwrapping from the other side, but in contrast to force-induced unwrapping, we find only a weak sequence-dependent asymmetry. Notably, centromeric CENP-A nucleosomes do not unwrap anti-cooperatively, in stark contrast to H3 nucleosomes. Finally, our results reconcile previous conflicting findings about the differences in height between H3 and CENP-A nucleosomes. We expect our approach to enable critical insights into epigenetic regulation of nucleosome structure and stability and to facilitate future high-throughput AFM studies that involve heterogeneous nucleoprotein complexes.

• Dokumententyp: Zeitschriftenartikel
• Fakultät: Physik
• Themengebiete: 500 Naturwissenschaften und Mathematik > 530 Physik
• ISSN: 2040-3364
• Sprache: Englisch