Nanoscale analysis of human G1 and metaphase chromatinin situ

Abstract

The structure of chromatin at the nucleosome level inside cells is mysterious. Here we present in situ cryo-ET analyses of chromatin in both G1 and metaphase RPE-1 cells. G1 nucleosomes are concentrated in globular chromatin domains and metaphase nucleosomes are concentrated in the chromatids. Classification analysis reveals that canonical mononucleosomes, ordered stacked dinucleosomes, and mononucleosomes with a disordered gyre-proximal density are abundant in both cell-cycle states. Class averages that have more than two stacked nucleosomes or that have side-by-side dinucleosomes are not detected, suggesting that groups of more than two nucleosomes are heterogeneous. Large multi-megadalton structures are abundant in G1 nucleoplasm, but not found in G1 chromatin domains and metaphase chromatin. The macromolecular phenotypes studied here represent a starting point for the comparative analysis of condensation in normal and unhealthy human cells, in other cell-cycle states, other organisms, and in vitro chromatin assemblies.

Hypothesis: The opposing pulling forces exerted by spindle microtubules can cause sliding of chromatin layers and facilitate sister chromatid resolution

“…Consistent with results indicating that chromosomes are helical structures (see above), it was proposed that the successive chromatin layers are connected forming a continuous helicoid (Daban, 2014) containing ~0.5 Mb per turn in human chromosomes. Since the long nested loops considered in Hi-C experiments presented in the preceding paragraph must be highly packed to achieve the high nucleosome density of metaphase chromosomes (Daban, 2003;Eltsov et al, 2008;Ou et al, 2017;Chen et al, 2023), it was suggested that they could be compacted into chromatin layers . Other authors (Grigoryev et al, 2016;Bascom and Schlick, 2017) suggested a hierarchical layering of loops to integrate the models based on a highly disordered chromatin filaments (Eltsov et al, 2008;Nishino et al, 2012) and results indicating a multilayer organization of chromatin in chromosomes.…”