Formation of cellular close-ended tunneling nanotubes through mechanical deformation.

Membrane nanotubes or tunneling nanotubes (TNTs) that connect cells have been recognized as a previously unidentified pathway for intercellular transport between distant cells. However, it is unknown how this delicate structure, which extends over tens of micrometers and remains robust for hours, is formed. Here, we found that a TNT develops from a double filopodial bridge (DFB) created by the physical contact of two filopodia through helical deformation of the DFB. The transition of a DFB to a close-ended TNT is most likely triggered by disruption of the adhesion of two filopodia by mechanical energy accumulated in a twisted DFB when one of the DFB ends is firmly attached through intercellular cadherin-cadherin interactions. These studies pinpoint the mechanistic questions about TNTs and elucidate a formation mechanism.

Anomalous diffusion of active Brownian particles cross-linked to a networked polymer: Langevin dynamics simulation and theory.

Quantitatively understanding the dynamics of an active Brownian particle (ABP) interacting with a viscoelastic polymer environment is a scientific challenge. It is intimately related to several interdisciplinary topics such as the microrheology of active colloids in a polymer matrix and the athermal dynamics of the in vivo chromosomes or cytoskeletal networks. Based on Langevin dynamics simulation and analytic theory, here we explore such a viscoelastic active system in depth using a star polymer of functionality f with the center cross-linker particle being ABP. We observe that the ABP cross-linker, despite its self-propelled movement, attains an active subdiffusion with the scaling ΔR2(t) ∼ tα with α ≤ 1/2, through the viscoelastic feedback from the polymer. Counter-intuitively, the apparent anomaly exponent α becomes smaller as the ABP is driven by a larger propulsion velocity, but is independent of functionality f or the boundary conditions of the polymer. We set forth an exact theory and show that the motion of the active cross-linker is a Gaussian non-Markovian process characterized by two distinct power-law displacement correlations. At a moderate Péclet number, it seemingly behaves as fractional Brownian motion with a Hurst exponent H = α/2, whereas, at a high Péclet number, the self-propelled noise in the polymer environment leads to a logarithmic growth of the mean squared displacement (∼ln t) and a velocity autocorrelation decaying as -t-2. We demonstrate that the anomalous diffusion of the active cross-linker is precisely described by a fractional Langevin equation with two distinct random noises.

Two conformational states in D-shaped DNA: Effects of local denaturation.

The bending of double-stranded(ds) DNA on the nano-meter scale plays a key role in many cellular processes such as nucleosome packing, transcription-control, and viral-genome packing. In our recent study, a nanometer-sized dsDNA bent into a D shape was formed by hybridizing a circular single-stranded(ss) DNA and a complementary linear ssDNA. Our fluorescence resonance energy transfer (FRET) measurement of D-DNA revealed two types of conformational states: a less-bent state and a kinked state, which can transform into each other. To understand the origin of the two deformed states of D-DNA, here we study the presence of open base-pairs in the ds portion by using the breathing-DNA model to simulate the system. We provide strong evidence that the two states are due to the emergence of local denaturation, i.e., a bubble in the middle and two forks at ends of the dsDNA portion. We also study the system analytically and find that the free-energy landscape is bistable with two minima representative of the two states. The kink and fork sizes estimated by the analytical calculation are also in excellent agreement with the results of the simulation. Thus, this combined experimental-simulation-analytical study corroborates that highly bent D-DNA reduces bending stress via local denaturation.

Dynamic Release of Bending Stress in Short dsDNA by Formation of a Kink and Forks.

Bending with high curvature is one of the major mechanical properties of double-stranded DNA (dsDNA) that is essential for its biological functions. The emergence of a kink arising from local melting in the middle of dsDNA has been suggested as a mechanism of releasing the energy cost of bending. Herein, we report that strong bending induces two types of short dsDNA deformations, induced by two types of local melting, namely, a kink in the middle and forks at the ends, which we demonstrate using D-shaped DNA nanostructures. The two types of deformed dsDNA structures dynamically interconvert on a millisecond timescale. The transition from a fork to a kink is dominated by entropic contribution (anti-Arrhenius behavior), while the transition from a kink to a fork is dominated by enthalpic contributions. The presence of mismatches in dsDNA accelerates kink formation, and the transition from a kink to a fork is removed when the mismatch size is three base pairs.

How a short double-stranded DNA bends.

A recent experiment using fluorescence microscopy showed that double-stranded DNA fragments shorter than 100 base pairs loop with the probabilities higher by the factor of 10(2)-10(6) than predicted by the worm-like chain (WLC) model [R. Vafabakhsh and T. Ha, Science 337, 1101(2012)]. Furthermore, the looping probabilities were found to be nearly independent of the loop size. The results signify a breakdown of the WLC model for DNA mechanics which works well on long length scales and calls for fundamental understanding for stressed DNA on shorter length scales. We develop an analytical, statistical mechanical model to investigate what emerges to the short DNA under a tight bending. A bending above a critical level initiates nucleation of a thermally induced bubble, which could be trapped for a long time, in contrast to the bubbles in both free and uniformly bent DNAs, which are either transient or unstable. The trapped bubble is none other than the previously hypothesized kink, which releases the bending energy more easily as the contour length decreases. It leads to tremendous enhancement of the cyclization probabilities, in a reasonable agreement with experiment.

Enhanced bubble formation in looped short double-stranded DNA.

Recent experiments have shown the double-stranded (ds) DNAs readily bend and loop over the scale much shorter than their persistence length (50 nm). In an effort to unveil this seemingly surprising phenomenon, we study the emergence of bubbles in short ds DNA loops by simulating the breathing DNA model. We analyze the bubble size distributions and the melting curves for varying contour lengths, which are critically compared with those of linear DNA of the same lengths. We analytically evaluate the free energies associated with double-strand bending and single-strand bubble formation to explain the simulation data. It is found that in shorter looped DNA the bubbles are more easily initiated and formed to release the large bending energy, giving rise to melting at a lower temperature and a lower contour length.

How double-stranded DNA breathing enhances its flexibility and instability on short length scales.

We study the unexpected high flexibility of short dsDNA which recently has been reported by a number of experiments. Via the Langevin dynamics simulation of our Breathing DNA model, first we observe the formation of bubbles within the duplex and also forks at the ends, with the size distributions independent of the contour length. We find that these local denaturations at a physiological temperature, despite their rare and transient presence, can lower the persistence length drastically for a short DNA segment in agreement with experiment.