Gradient Nanoconfinement Facilitates Binding of Transcriptional Factor NF-κB to Histone- and Protamine-DNA Complexes.

Mechanically induced chromosome reorganization plays important roles in transcriptional regulation. However, the interplay between chromosome reorganization and transcription activities is complicated, such that it is difficult to decipher the regulatory effects of intranuclear geometrical cues. Here, we simplify the system by introducing DNA, packaging proteins (i.e., histone and protamine), and transcription factor NF-κB into a well-defined fluidic chip with changing spatical confinement ranging from 100 to 500 nm. It is uncovered that strong nanoconfinement suppresses higher-order folding of histone- and protamine-DNA complexes, the fracture of which exposes buried DNA segments and causes increased quantities of NF-κB binding to the DNA chain. Overall, these results reveal a pathway of how intranuclear geometrical cues alter the open/closed state of a DNA-protein complex and therefore affect transcription activities: i.e., NF-κB binding.

An optically fabricated gradient nanochannel array to access the translocation dynamics of T4-phage DNA through nanoconfinement.

It has been widely recognized that nanostructures in natural biological materials play important roles in regulating life machinery. Even though nanofabrication techniques such as two-photon polymerization (TPP) provide sub-100 nm fabrication resolution, it remains technologically challenging to produce 3D nanoscale features modeling the complexity in vivo. We herein demonstrate that a nanochannel array carrying different sizes and nanostructures with gradually transitioning dimensions can be easily produced on a slightly tilted nano-stage. Using the gradient nanochannel array, we systematically investigate the factors affecting the dynamics of DNA translocation through nanoconfinement, including the size of biomolecules and geometrical features of the physical environment, which resembles the selectivity of nanopores in the cell membrane. It is observed that T4-phage DNA shows distinctive conformational transition dynamics during translocation through nanochannels driven by electric field or flow, and the deformation energy required for DNA to enter the nanochannels depends on both chemical environmental conditions, i.e., the ionic strength regulating DNA persistence length and nanochannel dimension. In the electric field, DNA repeatedly gets stretched and compressed during its migration through the nanochannel, reflected by elevated fluctuation in extension, which is substantially greater than the thermal fluctuation. However, driven by flow, DNA remains stretched during translocation through nanochannels, and shows variances in extension of merely a few hundred nanometers. These results indicate that the optically fabricated gradient nanochannel array is a suitable platform for optimizing the experimental conditions for biomedical applications such as gene mapping, and verify that production of complex three dimensional (3D) nanostructures can be greatly simplified by including slight inclination during TPP fabrication.

Generation and Modulation of Controllable Multi-Focus Array Based on Phase Segmentation.

A Circular-Sectorial Phase Segmentation (CSPS) noniterative method for effectively generating and manipulating muti-focus array (MFA) was proposed in this work. The theoretical model of the CSPS was built up based on vectorial diffraction integral and the phase modulation factor was deduced with inverse fast Fourier transform. By segmenting the entrance pupil into specified regions, which were sequentially assigned with the values carried out by phase modulation factor, the methodology could generate flexible MFAs with desired position and morphology. Subsequently, the CSPS was investigated in parallelized fabrication with a laser direct writing system. The positioning accuracy was greater than 96% and the morphologic consistency of the parallelly fabricated results was greater than 92%.