Scientists have discovered that for a long time the twisted structure of DNA, which was mistaken for a knot, is actually something entirely different.
Within cells, DNA is twisted, copied and separated. Twist can affect gene function and affect how it can be switched. Studying how DNA responds to stress can help scientists better understand how genes are regulated, how molecules are organized, and how problems with these processes contribute to disease.
For years, researchers have used nanopores (small holes wide enough for a single DNA strand to slip through), and have read DNA sequences quickly and cheaply. These systems work by measuring the current flowing through the nanopore. When a DNA molecule passes, it destroys its current in a clear way that corresponds to each of the four “characters” that make up the code of DNA: a, t, c, g.
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Unexpected deceleration or spikes of this signal were often interpreted as DNA knots. However, a new study now published on August 12th in Journal Physics Review X shows that changes in these signals also mean plectonem.
“Knots and plectnems can be very similar in nanopore signals,” Ulrich Keeser, a physicist at the Cavendish Institute at Cambridge University, told Live Science. “But they come from very different physical mechanisms. The knot is like a tight entanglement. The plectonem is similar to a coiled spring formed by torque.”
To study these coils, researchers passed DNA strands through corn-shaped nanopores in salty solutions with high pH. This solution helped create an electroosmotic flow. In other words, it started to spin when DNA entered the pores. The movement produced sufficiently strong torsional force or torque wrapped around the DNA, explained Keiser.
Related: DNA has an expiration date. But proteins reveal secrets about our ancient ancestors that we never thought of.
Keyser and his team applied voltage across the nanopore to drive DNA and measure changes in current.
“In these types of nanoscale systems, everything is very high friction, so DNA moves like it’s swimming through honey,” Keeser said. “This is a very viscous environment, so this corkscrew moves to push DNA and it’s a relatively high force.”
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Researchers analyzed thousands of events from these events. Although some knots still appeared in the experiment, they tended to be smaller – about 140 nanometers in diameter – the plectonem was about 2,100 nanometers. As the voltage applied to the system increases, plectnems have become more common due to the stronger torque.
To further test how twists affect DNA behavior, researchers introduced a small break called Nicks into one strand of the double helix of DNA. These nicks made it possible to rotate the DNA more easily and release the built-in tension. This confirmed that torsional stress is an important factor in the formation of these structures.
“Controlling the ability of molecules to rotate allowed us to change the frequency with which plectonems appeared,” Keeser said.
Although nanopores are very different from living cells, these types of plectonems can also form during processes such as DNA transcription and replication. Transcription explains that the code for DNA is copied by another molecule called RNA and shipped to a cell. Replication describes when DNA molecules are fully replicated. This happens when, for example, cells divide.
“We believe that molecular twists can actually cause the formation of i-motifs and g-quadruplexes,” Keyser told Live Science, naming two specific knots found in DNA. Therefore, he explained that what they found in their lab studies is likely to affect living cells.
Keyser and his team have been investigating how plectonems and other DNA structures form during natural processes such as transcription. Previous studies investigated how torsional stress affects DNA replication. Nanopores give scientists a way to not only read DNA, but also see how it behaves, this study emphasizes.
“It’s just the fact that a DNA molecule can squeeze pores that are assumed to be much larger than the pore size,” Slaven Garage, a physicist at the National University of Singapore who is not part of the study, told Live Science. “It’s 10, 50, and it’s 100 times harder than the size of the pores. It still bends and passes through.”
Garage was excited by the findings. In the future, “the nanopore-induced torsion may be able to be isolated from the torsion that had previously fallen into DNA. This is important to understand how coils and knots regulate gene activity.
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