Chromatin Is Frequently Unknotted at the Megabase Scale

Abstract

Knots in the human genome would greatly impact diverse cellular processes ranging from transcription to gene regulation. To date, it has not been possible to directly examine the genome in vivo for the presence of knots. Recently, methods for serial fluorescent in situ hybridization have made it possible to measure the three-dimensional position of dozens of consecutive genomic loci in vivo. However, the determination of whether genomic trajectories are knotted remains challenging because small errors in the localization of a single locus can transform an unknotted trajectory into a highly knotted trajectory and vice versa. Here, we use stochastic closure analysis to determine if a genomic trajectory is knotted in the setting of experimental noise. We analyze 4727 deposited genomic trajectories of a 2-Mb-long chromatin interval from human chromosome 21. For 243 of these trajectories, their knottedness could be reliably determined despite the possibility of localization errors. Strikingly, in each of these 243 cases, the trajectory was unknotted. We note a potential source of bias insofar as knotted contours may be more difficult to reliably resolve. Nevertheless, our data are consistent with a model in which, at the scales probed, the human genome is often free of knots.

Exploring chromosomal structural heterogeneity across multiple cell lines

“…The heterogeneity has now been confirmed by direct imaging of individual chromosomal structures ( Bintu et al, 2018 ; Boettiger et al, 2016 ; Nir et al, 2018 ). As a consequence of this conformational plasticity, statistical ensembles ( Zhang and Wolynes, 2015 ; Di Pierro et al, 2016 ; Di Pierro et al, 2017 ; Di Pierro et al, 2018 ; Zhang and Wolynes, 2016 ; Di Pierro, 2019 ; Goundaroulis et al, 2020 ; Bascom et al, 2019 ; Dekker et al, 2013 ; Kalhor et al, 2012 ) must be used in order to describe chromosomal structures in vivo.…”

Linker histone H1.8 inhibits chromatin binding of condensins and DNA topoisomerase II to tune chromosome length and individualization

“…Replicated pairs of chromatids are linked by cohesin during interphase, and cohesin removal during prophase (except at centromeres and other limited protected loci) promotes resolution of sister chromatids, together with actions of condensin II and topo II ( Nagasaka et al, 2016 ; Gandhi et al, 2006 ). Different chromosomes are also largely unentangled in interphase HeLa cells ( Goundaroulis et al, 2020 ; Tavares-Cadete et al, 2020 ), and Ki-67 localization on chromosome peripheries may act as a steric and electrostatic barrier to prevent interchromosomal entanglement during mitosis ( Cuylen et al, 2016 ). However, some interchromosomal linkages were observed in metaphase ( Potapova et al, 2019 ; Marko, 2008 ).…”

Parameter-free molecular super-structures quantification in single-molecule localization microscopy

“…Additionally, recent super-resolution studies indicate that chromatin is also functionally organized in connected nano-scale compartments (Prakash et al, 2015;Szabo et al, 2018;Nir et al, 2018;Maiser ............................................................................................................................................................................. et al, 2020). Rapidly evolving methods of chromatin tracing (Boettiger et al, 2016;Wang et al, 2016;Beliveau et al, 2015;Nir et al, 2018;Bintu et al, 2018) and super-resolved imaging of the accessible genome (Xie et al, 2020) require sophisticated algorithms to analyze the topology of the generated paths (Goundaroulis et al, 2020). To understand the relationship between these complex structures and the underlying biological mechanism and functions of the genome (Bronshtein et al, 2015;Khanna et al, 2019;Leidescher et al, 2020 Preprint;Smeets et al, 2014), a more sophisticated and standardized analysis of SMLM data is urgently required.…”