Chromatinization modulates topoisomerase II processivity
AbstractType IIA topoisomerases are essential DNA processing enzymes that must robustly and reliably relax DNA torsional stress. While cellular processes constantly create varying torsional stress, how this variation impacts type IIA topoisomerase function remains obscure. Using multiple single-molecule approaches, we examined the torsional dependence of eukaryotic topoisomerase II (topo II) activity on naked DNA and chromatin. We observed that topo II is ~50-fold more processive on buckled DNA than previously estimated.
Etoposide promotes DNA loop trapping and barrier formation by topoisomerase II
Etoposide is a broadly employed chemotherapeutic and eukaryotic topoisomerase II poison that stabilizes cleaved DNA intermediates to promote DNA breakage and cytotoxicity. How etoposide perturbs topoisomerase dynamics is not known. Here we investigated the action of etoposide on yeast topoisomerase II, human topoisomerase IIÎ± and human topoisomerase IIÎ² using several sensitive single-molecule detection methods. Unexpectedly, we found that etoposide induces topoisomerase to trap DNA loops, compacting DNA and restructuring DNA topology.
Polarity of the CRISPR roadblock to transcription
CRISPR (clustered regularly interspaced short palindromic repeats) utility relies on a stable Cas effector complex binding to its target site. However, a Cas complex bound to DNA may be removed by motor proteins carrying out host processes and the mechanism governing this removal remains unclear. Intriguingly, during CRISPR interference, RNA polymerase (RNAP) progression is only fully blocked by a bound endonuclease-deficient Cas (dCas) from the protospacer adjacent motif (PAM)-proximal side.
Resonator nanophotonic standing-wave array trap for single-molecule manipulation and measurement
Nanophotonic tweezers represent emerging platforms with significant potential for parallel manipulation and measurements of single biological molecules on-chip. However, trapping force generation represents a substantial obstacle for their broader utility. Here, we present a resonator nanophotonic standing-wave array trap (resonator-nSWAT) that demonstrates significant force enhancement. This platform integrates a critically-coupled resonator design to the nSWAT and incorporates a novel trap reset scheme.
Angular Optical Trapping to Directly Measure DNA Torsional Mechanics
Angular optical trapping (AOT) is a powerful technique that permits direct angular manipulation of a trapped particle with simultaneous measurement of torque and rotation, while also retaining the capabilities of position and force detection. This technique provides unique approaches to investigate the torsional properties of nucleic acids and DNA-protein complexes, as well as impacts of torsional stress on fundamental biological processes, such as transcription and replication.
Torsional Stiffness of Extended and Plectonemic DNA
DNA torsional elastic properties play a crucial role in DNA structure, topology, and the regulation of motor protein progression. However, direct measurements of these parameters are experimentally challenging. Here, we present a constant-extension method integrated into an angular optical trap to directly measure torque during DNA supercoiling. We measured the twist persistence length of extended DNA to be 22 nm under an extremely low force (âˆ¼0.02 pN) and the twist persistence length of plectonemic DNA to be 24 nm.
Optical tweezers in single-molecule biophysics
Optical tweezers have become the method of choice in single-molecule manipulation studies. In this Primer, we first review the physical principles of optical tweezers and the characteristics that make them a powerful tool to investigate single molecules. We then introduce the modifications of the method to extend the measurement of forces and displacements to torques and angles, and to develop optical tweezers with single-molecule fluorescence detection capabilities.
Dextran-coated iron oxide nanoparticle-induced nanotoxicity in neuron cultures
Recent technological advances have introduced diverse engineered nanoparticles (ENPs) into our air, water, medicine, cosmetics, clothing, and food. However, the health and environmental effects of these increasingly common ENPs are still not well understood. In particular, potential neurological effects are one of the most poorly understood areas of nanoparticle toxicology (nanotoxicology), in that low-to-moderate neurotoxicity can be subtle and difficult to measure.
Towards biological applications of nanophotonic tweezers
Optical trapping (synonymous with optical tweezers) has become a core biophysical technique widely used for interrogating fundamental biological processes on size scales ranging from the single-molecule to the cellular level. Recent advances in nanotechnology have led to the development of â€˜nanophotonic tweezers,â€™ an exciting new class of â€˜on-chipâ€™ optical traps. Here, we describe how nanophotonic tweezers are making optical trap technology more broadly accessible and bringing unique biosensing and manipulation capabilities to biological applications of optical trapping.