Dr Dominika Gruszka

Abstract:

Biological processes are orchestrated by complex networks of molecules. Conventional approaches for studying the action of biomolecules operate on a population level, averaging out any inhomogeneities within the ensemble. Investigating one biological macromolecule at a time allows researchers to directly probe individual behaviours, and thus characterise the intrinsic molecular heterogeneity of the system. Single-molecule methods have unravelled unexpected modes of action for many seemingly well-characterised biomolecules and often proved instrumental in understanding the intricate mechanistic basis of biological processes. This collection of reviews aims to showcase how single-molecule techniques can be used to address important biological questions and to inspire biochemists to ‘zoom in’ to the population and probe individual molecular behaviours, beyond the ensemble average. Furthermore, this issue of Essays in Biochemistry is the very first written and edited entirely by early career researchers, and so it also highlights the strength, diversity and excellence of the younger generation single-molecule scientists who drive this exciting field of research forward.

Abstract:

During replication, nucleosomes are disrupted ahead of the replication fork, followed by their reassembly on daughter strands from the pool of recycled parental and new histones. However, because no previous studies have managed to capture the moment that replication forks encounter nucleosomes, the mechanism of recycling has remained unclear. Here, through real-time single-molecule visualization of replication fork progression in Xenopus egg extracts, we determine explicitly the outcome of fork collisions with nucleosomes. Most of the parental histones are evicted from the DNA, with histone recycling, nucleosome sliding, and replication fork stalling also occurring but at lower frequencies. Critically, we find that local histone recycling becomes dominant upon depletion of endogenous histones from extracts, revealing that free histone concentration is a key modulator of parental histone dynamics at the replication fork. The mechanistic details revealed by these studies have major implications for our understanding of epigenetic inheritance.