How molecular motors give chromosomes their structure

As well as being an information processor, the genome is also a physical object, inspiring researchers to explore how its structure determines its function.

Fundamental Research

Health

Some 150 years ago, Walther Flemming studied chromosomal changes during cell division. Yet despite subsequent advances in knowledge, researchers have only recently identified the key elements that determine the organisation of chromosomes in the nucleus. “Chromosomes are fundamentally structured in DNA loops, with ring-shaped proteins known as structural maintenance of chromosomes (SMC) complexes, implicated in their creation,” says Cees Dekker(opens in new window), physics professor at Delft University of Technology(opens in new window) in the Netherlands and principal investigator of LoopingDNA. “The exact mechanisms for how these loops were formed and controlled however, have remained mysterious.” In 2018, Dekker’s lab discovered that an SMC called ‘condensin’ acts as a molecular motor(opens in new window) that loops DNA, giving chromosomes their structure. Now, using single-molecule assays, the LoopingDNA project has revealed more about the mechanisms involved and explored the impact of chromosome structure on biology.

A novel class of DNA motors

Supported by the European Research Council(opens in new window) (ERC), the LoopingDNA team used time-lapse single-molecule imaging to follow the condensin-driven DNA looping process in action. “We put a piece of DNA on a glass slide, visualised it, then saw the loop appearing over time as SMCs attached to the DNA,” explains Dekker. “Further experiments enabled us to observe many additional features, such as looping asymmetry and direction changes.” Additionally, the team discovered a new loop motif, called z-loops(opens in new window), formed when two loop-extruding condensin SMCs encounter and pass each other, shedding light on how DNA is organised into a tightly compacted structure. Loop extrusion was also found to intricately connect to the ‘supercoiling’ of DNA, which increases the amount of twist in a DNA strand. “We discovered that SMCs do not merely extrude DNA, but also add a twist with each extrusion, implying DNA supercoiling is regulated by the genome,” adds Dekker.

Mechanisms underlying DNA looping

Other fascinating insights have been gained by the project. Cellular DNA is covered in DNA-binding proteins such as nucleosomes or RNA polymerase, that could theoretically impede extrusion. The team were surprised to find this was not the case – SMCs are seemingly capable of subsuming virtually all objects into their DNA loop. This is significant, as proteins on DNA can be up to tens of nanometres in size, and the ring-shaped SMC is around 30 nanometres across. Dekker found that the ring could even accommodate DNA-bound gold particles 200 nanometres in size, larger than the ring of the extruding SMC itself. “We concluded that the DNA was not, as previously hypothesised, topologically contained within the SMC complex ring,” says Dekker. “We have now worked out a model whereby binding with adenosine triphosphate changes the SMC’s shape and size, bringing us closer to understanding the motor mechanism underlying DNA looping.”

Insights into genetic disorders

The researchers also set themselves the ambitious task of building a chromosome from scratch. Starting with a length of bare DNA (an Escherichia coli bacterial genome stripped of proteins), the aim was to add protein complexes, and other DNA-processing proteins, to study the effects of DNA-organising elements. While they achieved a basic version, DNA’s extreme fragility rendered a fully fledged version beyond the project’s resources. “If we can overcome these technical hurdles, our ‘genome-in-a-box’ could one day offer a toolkit for experiments that reveal how chromosomal structure regulates gene expression, offering insights into genetic disorders,” explains Dekker. With condensin known to be vital for healthy embryonic development, Dekker’s new consortium, currently pursuing funding through an ERC synergy grant, will research this area further. Of particular interest is how DNA looping may enable so-called ‘enhancers’ to get closer to the promoters of key genes, boosting their expression.