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Monday, February 20, 2017

Single molecule force spectroscopy reveals the effect of BiP chaperone on protein folding

María Paz Ramírez, Maira Rivera, Diego Quiroga-Roger, Andrés Bustamante, Marcela Vega, Mauricio Baez, Elias M. Puchner, Christian A.M. Wilson

BiP (Immunoglobulin Binding Protein) is a member of the Hsp70 chaperones that participates in protein folding in the endoplasmic reticulum. The function of BiP relies on cycles of ATP hydrolysis driving the binding and release of its substrate proteins. It still remains unknown how BiP affects the protein folding pathway and there has been no direct demonstration showing which folding state of the substrate protein is bound by BiP, as previous work has used only peptides. Here, we employ optical tweezers for single molecule force spectroscopy experiments to investigate how BiP affects the folding mechanism of a complete protein and how this effect depends on nucleotides. Using the protein MJ0366 as the substrate for BiP, we performed pulling and relaxing cycles at constant velocity to unfold and refold the substrate. In the absence of BiP, MJ0366 unfolded and refolded in every cycle. However, when BiP was added, the frequency of folding events of MJ0366 significantly decreased, and the loss of folding always occurred after a successful unfolding event. This process was dependent on ATP and ADP, since when either ATP was decreased or ADP was added, the duration of periods without folding events increased. Our results show that the affinity of BiP for the substrate protein increased in these conditions, which correlates with previous studies in bulk. Therefore, we conclude that BiP binds to the unfolded state of MJ0366 and prevents its refolding, and that this effect is dependent on both the type and concentration of nucleotides.

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Holographic optical tweezers-based in vivo manipulations in zebrafish embryos

Florian Hörner, Robert Meissner, Sruthi Polali, Jana Pfeiffer, Timo Betz, Cornelia Denz, Erez Raz

Understanding embryonic development requires the characterization of the forces and the mechanical features that shape cells and tissues within the organism. In addition, experimental application of forces on cells and altering cell and organelle shape allows determining the role such forces play in morphogenesis. Here, we present a holographic optical tweezers-based new microscopic platform for in vivo applications in the context of a developing vertebrate embryo that unlike currently used setups allows simultaneous trapping of multiple objects and rapid comparisons of viscoelastic properties in different locations. This non-invasive technique facilitates a dynamic analysis of mechanical properties of cells and tissues without intervening with embryonic development. We demonstrate the application of this platform for manipulating organelle shape and for characterizing the mechanobiological properties of cells in live zebrafish embryos. The method of holographic optical tweezers as described here is of general interest and can be easily transferred to studying a range of developmental processes in zebrafish, thereby establishing a versatile platform for similar investigations in other organisms.

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Microfluidics cell sample preparation for analysis: Advances in efficient cell enrichment and precise single cell capture

Liang Huang, Shengtai Bian, Yinuo Cheng, Guanya Shi, Peng Liu, Xiongying Ye, Wenhui Wang

Single cell analysis has received increasing attention recently in both academia and clinics, and there is an urgent need for effective upstream cell sample preparation. Two extremely challenging tasks in cell sample preparation—high-efficiency cell enrichment and precise single cell capture—have now entered into an era full of exciting technological advances, which are mostly enabled by microfluidics. In this review, we summarize the category of technologies that provide new solutions and creative insights into the two tasks of cell manipulation, with a focus on the latest development in the recent five years by highlighting the representative works. By doing so, we aim both to outline the framework and to showcase example applications of each task. In most cases for cell enrichment, we take circulating tumor cells (CTCs) as the target cells because of their research and clinical importance in cancer. For single cell capture, we review related technologies for many kinds of target cells because the technologies are supposed to be more universal to all cells rather than CTCs. Most of the mentioned technologies can be used for both cell enrichment and precise single cell capture. Each technology has its own advantages and specific challenges, which provide opportunities for researchers in their own area. Overall, these technologies have shown great promise and now evolve into real clinical applications.

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Elastic back-scattering patterns via particle surface roughness and orientation from single trapped airborne aerosol particles

Richard Fu, Chuji Wang, Olga Muñoz, Gorden Videen, Joshua L. Santarpia, Yong-Le Pan

We demonstrate a method for simultaneously measuring the back-scattering patterns and images of single laser-trapped airborne aerosol particles. This arrangement allows us to observe how the back-scattering patterns change with particle size, shape, surface roughness, orientation, etc. The recoded scattering patterns cover the angular ranges of θ=167.7–180° (including at 180° exactly) and ϕ=0–360° in spherical coordinates. The patterns show that the width of the average speckle intensity islands or rings is inversely proportional to particle size and how the shape of these intensity rings or islands also depends on the surface roughness. For an irregularly shaped particle with substantial roughness, the back-scattering patterns are formed with speckle intensity islands, the size and orientations of these islands depend more on the overall particle size and orientation, but have less relevance to the fine alteration of the surface structure and shapes. The back-scattering intensity at 180° is very sensitive to the particle parameters. It can change from a maximum to a minimum with a change of 0.1% in particle size or refractive index. The method has potential use in characterizing airborne aerosol particles, and may be used to provide back-scattering information for LIDAR applications.

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The role of cyclic di-GMP and exopolysaccharide in type IV pilus dynamics

Jan Ribbe, Amy E. Baker, Sebstian Euler, George A. O'Toole and Berenike Maier

For Pseudomonas aeruginosa the levels of cyclic di-GMP govern the transition from the planktonic state to biofilm formation. Type IV pili (T4P) are crucial determinants of biofilm structure and dynamics, but it is unknown how the levels of c-di-GMP affect pilus dynamics. Here, we scrutinized how c-di-GMP affects molecular motor properties and adhesive behavior of T4P. By means of retraction, T4P generated forces of ∼ 30 pN. Deletion mutants in the proteins with known roles in biofilm formation, swarming motility and exopolysaccharide (EPS) production, specifically, the diguanulate cyclases sadC and roeA or the c-di-GMP phosphodiesterase bifA, showed only modest effects on velocity or force of T4P retraction. At high levels of c-di-GMP, the production of exopolysaccharides (EPS) and in particular of Pel is upregulated. We found that Pel production strongly enhances T4P-mediated surface adhesion of P. aeruginosa, suggesting that that T4P — matrix interactions may be involved in biofilm formation by P. aeruginosa. Finally, our data are consistent with the previously proposed sling-shot-like “twitching” motility of P. aeruginosa.

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Friday, February 17, 2017

Measurement of laterally induced optical forces at the nanoscale

Fei Huang, Venkata Ananth Tamma, Mohsen Rajaei, Mohammad Almajhadi, and H. Kumar Wickramasinghe

We demonstrate the measurement of laterally induced optical forces using an Atomic Force Microscope (AFM). The lateral electric field distribution between a gold coated AFM probe and a single nano-aperture in a gold film is mapped by measuring the lateral optical force between the apex of the AFM probe and the nano-aperture. The fundamental torsional eigen-mode of an AFM cantilever probe was used to detect the laterally induced optical forces. We engineered the cantilever shape using focused ion beam milling to improve the detected signal to noise ratio. The measured distributions of lateral optical force agree well with electromagnetic simulations of the metal coated AFM probe interacting with the nano-aperture. This technique can be extended to simultaneously detect both lateral and longitudinal optical forces at the nanoscale by using an AFM cantilever as a multi-channel detector. This will enable simultaneous Photon Induced Force Microscopy detection of molecular responses with different incident field polarizations. The technique can be implemented on both cantilever and tuning fork based AFMs.

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Force-Dependent Folding and Unfolding Kinetics in DNA Hairpins Reveals Transition-State Displacements along a Single Pathway

Anna Alemany and Felix Ritort

Biomolecules diffusively explore their energy landscape overcoming energy barriers via thermally activated processes to reach the biologically relevant conformation. Mechanically induced unfolding and folding reactions offer an excellent playground to feature these processes at the single-molecule level by monitoring changes in the molecular extension. Here we investigate two-state DNA hairpins designed to have the transition states at different locations. We use optical tweezers to characterize the force-dependent behavior of the kinetic barrier from nonequilibrium pulling experiments by using the continuous effective barrier approach (CEBA). We introduce the mechanical fragility and the molecular transition-state susceptibility, both useful quantities to characterize the response of the transition state to an applied force. Our results demonstrate the validity of the Leffler–Hammond postulate where the transition state approaches the folded state as force increases, implying monotonically decreasing fragility with force and a non-negative transition state susceptibility at all forces.

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Photon-phonon Interaction in a Microfiber Induced by Optical and Electrostrictive Forces

Yun-chao Shi, Wei Luo, Fei Xu & Yan-qing Lu

Stimulated Brillouin scattering (SBS) via electrostrictive force is a fundamental interaction between light and sound which limits the power in conventional optical fibers. The emergence of optical microfibers with subwavelength diameter, ultralight mass and an intense light field, provides a new platform for photon–phonon coupling, resulting in the radiation pressure mediated contribution of SBS. This study examines the optomechanical system in cylindrical coordinates, reveals the theoretically radiation pressure induced analogous, and demonstrates contrary effect compared with electrostrictive force in solid or hollow silica microfibers. The finding shows that the photon-phonon coupling, which is related to SBS, can be suppressed in a solid microfiber, and even be completely cancelled in a hollow microfiber.

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Unfolding DNA condensates produced by DNA-like charged depletants: A force spectroscopy study

C. H. M. Lima, M. S. Rocha, and E. B. Ramos

In this work, we have measured, by means of optical tweezers, forces acting on depletion-induced DNA condensates due to the presence of the DNA-like charged protein bovine serum albumin (BSA). The stretching and unfolding measurements performed on the semi-flexible DNA chain reveal (1) the softening of the uncondensed DNA contour length and (2) a mechanical behavior strikingly different from those previously observed: the force-extension curves of BSA-induced DNA condensates lack the “saw-tooth” pattern and applied external forces as high as ≈80pN≈80 pN are unable to fully unfold the condensed DNA contour length. This last mechanical experimental finding is in agreement with force-induced “unpacking” detailed Langevin dynamics simulations recently performed by Cortini et al. on model rod-like shaped condensates. Furthermore, a simple thermodynamics analysis of the unfolding process has enabled us to estimate the free energy involved in the DNA condensation: the estimated depletion-induced interactions vary linearly with both the condensed DNA contour length and the BSA concentration, in agreement with the analytical and numerical analysis performed on model DNA condensates. We hope that future additional experiments can decide whether the rod-like morphology is the actual one we are dealing with (e.g. pulling experiments coupled with super-resolution fluorescence microscopy).

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Oscillations of absorbing particles at the water-air interface induced by laser tweezers

Min-Cheng Zhong, Zi-Qiang Wang, and Yin-Mei Li

We present an experimental study on oscillation of absorbing particles at the water-air interface. The oscillation is induced by laser tweezers, which are generated with a high numerical aperture objective. When the laser beam is tightly focused at the water-air interface, the optical gradient force attracts the particles to the spot center, and the laser heating of particles results in a strong thermal gradient that drives the particles to leave the spot center. Under the action of thermal and optical gradient force together, the absorbing particles oscillate at the water-air interface.

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Wednesday, February 15, 2017

Antibodies Damage the Resilience of Fimbriae, Causing Them To Be Stiff and Tangled

Bhupender Singh, Narges Mortezaei, Stephen J. Savarino, Bernt Eric Uhlin, Esther Bullitt and Magnus Andersson

As adhesion fimbriae are a major virulence factor for many pathogenic Gram-negative bacteria, they are also potential targets for antibodies. Fimbriae are commonly required for initiating the colonization that leads to disease, and their success as adhesion organelles lies in their ability to both initiate and sustain bacterial attachment to epithelial cells. The ability of fimbriae to unwind and rewind their helical filaments presumably reduces their detachment from tissue surfaces with the shear forces that accompany significant fluid flow. Therefore, the disruption of functional fimbriae by inhibiting this resilience should have high potential for use as a vaccine to prevent disease. In this study, we show that two characteristic biomechanical features of fimbrial resilience, namely, the extension force and the extension length, are significantly altered by the binding of antibodies to fimbriae. The fimbriae that were studied are normally expressed on enterotoxigenic Escherichia coli, which are a major cause of diarrheal disease. This alteration in biomechanical properties was observed with bivalent polyclonal antifimbrial antibodies that recognize major pilin subunits but not with the Fab fragments of these antibodies. Thus, we propose that the mechanism by which bound antibodies disrupt the uncoiling of natural fimbria under force is by clamping together layers of the helical filament, thereby increasing their stiffness and reducing their resilience during fluid flow. In addition, we propose that antibodies tangle fimbriae via bivalent binding, i.e., by binding to two individual fimbriae and linking them together. Use of antibodies to disrupt physical properties of fimbriae may be generally applicable to the large number of Gram-negative bacteria that rely on these surface-adhesion molecules as an essential virulence factor.

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Dynamics of polystyrene beads linking to DNA molecules under single optical tweezers: A numerical study using full normalized Langevin equation

Thai Dinh Trung, Bui Xuan Kien, Nguyen Thanh Tung, Ho Quang Quy

The dynamics of a polystyrene bead linking to a λλ-phage deoxyribonucleic acid (DNA) molecule under single optical tweezers using CW laser beam is analyzed by the full normalized Langevin equation with elastic and Brownian forces. The obtained results show that the stable time and stable position of polystyrene bead depend on its beginning position, laser beam waist, and laser beam power. The oscillation of polystyrene beads at stable position is observed with different parameters of optical tweezers. Finally, the conditions to stabilize the bead and to stretch the λλ-phage DNA by single laser beam optical tweezers are discussed.

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Optical tweezers affected by monochromatic aberrations

R. Kampmann and S. Sinzinger

The performance of optical tweezers in the presence of monochromatic aberrations is investigated theoretically and experimentally. For the calculation of optical forces we use a ray-optics-based force simulation, which is embedded in a ray tracing routine. By considering optical path differences caused by monochromatic aberrations, we are able to simulate their effect on the trapping forces. Optical trapping experiments considering the influence of aberrations were performed with an optical tweezers system, specially adapted to trap particles in gaseous surrounding. The conformity of these measurements with the theoretical predictions verifies the correct performance of the optical force simulation routine. Based on the reliable simulation tool, further theoretical studies are performed on how optical aberrations affect the function and performance of optical tweezers.

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Practical guide to the realization of a convertible optical trapping system

Chenglong Zhao
In this article, we provide a detailed guide to the construction of a convertible optical trapping system for either single-beam or counter-propagating trap. The single-beam trap maintains all the functionalities that a conventional optical tweezer has. While the counter-propagating trap allows for the trapping of particles that single-beam trap cannot handle. The counter-propagating trap can be easily switched to a single-beam trap, and vice versa. Therefore, this convertible optical trapping system allows for the trapping and manipulation of particles with a wide variety of sizes and materials.

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Influence of evanescent wave on birefringent microplates

O.V. Angelsky, S.G. Hanson, P.P. Maksimyak, A.P. Maksimyak, C.Yu. Zenkova, P.V. Polyanskii, and D.I. Ivanskyi

Mechanical action caused by the optical forces connected with the canonical momentum density associated with the local wavevector or Belinfante’s spin angular momentum is experimentally verified. The helicity-dependent and the helicity-independent forces determined by spin momenta of different nature open attractive prospects for the use of optical structures for manipulating minute quantities of matter of importance in nanophysics, nanooptics and nanotechnologies, precision chemistry and pharmacology and in numerous other areas. Investigations in this area reveal new, extraordinary manifestations of optical forces, including the helicity-independent force caused by the transverse helicity-independent spin or vertical spin of a diagonally polarized wave, which was not observed and exploited up to recently. The main finding of our study consists in a direct experimental demonstration of the physical existence and mechanical action of this recently discovered extraordinary transverse component of the spin here arising in an evanescent light wave due to the total internal reflection of a linearly polarized probing beam with azimuthal angle 45° at the interface between the birefringent plate and air, which is oriented perpendicularly to the wavevector of an evanescent wave and localized over the boundary of the transparent media with polarization-dependent refraction indices.

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Tuesday, February 14, 2017

Experimental measurement of binding energy, selectivity, and allostery using fluctuation theorems

Joan Camunas-Soler, Anna Alemany, Felix Ritort

Thermodynamic bulk measurements of binding reactions rely on the validity of the law of mass action and the assumption of a dilute solution. Yet, important biological systems such as allosteric ligand-receptor binding, macromolecular crowding, or misfolded molecules may not follow these assumptions and may require a particular reaction model. Here we introduce a fluctuation theorem for ligand binding and an experimental approach using single-molecule force spectroscopy to determine binding energies, selectivity, and allostery of nucleic acids and peptides in a model-independent fashion. A similar approach could be used for proteins. This work extends the use of fluctuation theorems beyond unimolecular folding reactions, bridging the thermodynamics of small systems and the basic laws of chemical equilibrium.

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Accurate Representations of the Physicochemical Properties of Atmospheric Aerosols: When are Laboratory Measurements of Value?

Aleksandra Marsh, Grazia Rovelli, Young-Chul Song, Kelly L. Pereira, Rose E. Willoughby, Bryan R. Bzdek, Jacqueline Hamilton, Andrew Orr-Ewing, David Owen Topping and Jonathan P Reid

Laboratory studies can provide important insights into the processes that occur at the mesoscale in ambient aerosol. We examine the accuracies of measurements of core physicochemical properties of aerosols that can be made in single particle studies and explore the impact of these properties on the microscopic processes that occur in ambient aerosol. Presenting new measurements, we examine here the refinements in our understanding of aerosol hygroscopicity, surface tension, viscosity and optical properties that can be gained from detailed laboratory measurements for complex mixtures through to surrogates for secondary organic atmospheric aerosols.

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DNA interaction with DAPI fluorescent dye: Force spectroscopy decouples two different binding modes

L. A. Reis, M. S. Rocha

In this work we use force spectroscopy in order to investigate the interaction between the DAPI fluorescent dye and the λ-DNA molecule under high (174 mM) and low (34 mM) ionic strengths. Firstly we have measured the changes on the mechanical properties (persistence and contour lengths) of the DNA-DAPI complexes as a function of the dye concentration in the sample. Then, we use recently developed models in order to connect the behavior of both mechanical properties to the physical chemistry of the interaction. Such analysis has allowed us to identify and to decouple two main binding modes, determining the relevant physicochemical (binding) parameters for each of these modes: minor groove binding, which saturates at very low DAPI concentrations (CT ∼ 0.50 μM) and presents equilibrium binding constants of the order of ∼ 107 M– 1 for the two ionic strengths studied; and intercalation, which starts to play a significant role only after the saturation of the first mode, presenting much smaller equilibrium binding constants (∼ 105 M– 1).

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Coalescence Sampling and Analysis of Aerosols using Aerosol Optical Tweezers

Allen E. Haddrell, Rachael E. H. Miles, Bryan R. Bzdek, Jonathan P. Reid, Rebecca J. Hopkins, and Jim S. Walker

We present a first exploratory study to assess the use of aerosol optical tweezers as an instrument for sampling and detecting accumulation- and coarse-mode aerosol. A subpicoliter aqueous aerosol droplet is captured in the optical trap and used as a sampling volume, accreting mass from a free-flowing aerosol generated by a medical nebulizer or atomizer. Real-time measurements of the initial stability in size, refractive index, and composition of the sampling droplet inferred from Raman spectroscopy confirm that these quantities can be measured with high accuracy and low noise. Typical standard deviations in size and refractive index of the sampling droplet over a period of 200 s are <±2 nm and <±0.0005, respectively, equivalent to <±0.04% in both measured quantities. A standard deviation of <±1% over a 200 s period is achieved in the spontaneous Raman intensity measurement. When sampling coarse-mode aerosol, mass changes of <10 pg can be detected by the sampling droplet as discrete coalescence events. With accumulation-mode aerosol, we show that fluxes as low as 0.068 pg s–1 can be detected over a 50 s period, equivalent to ∼3 pg of sampled material.

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Driven optical matter: Dynamics of electrodynamically coupled nanoparticles in an optical ring vortex

Patrick Figliozzi, Nishant Sule, Zijie Yan, Ying Bao, Stanislav Burov, Stephen K. Gray, Stuart A. Rice, Suriyanarayanan Vaikuntanathan, and Norbert F. Scherer

To date investigations of the dynamics of driven colloidal systems have focused on hydrodynamic interactions and often employ optical (laser) tweezers for manipulation. However, the optical fields that provide confinement and drive also result in electrodynamic interactions that are generally neglected. We address this issue with a detailed study of interparticle dynamics in an optical ring vortex trap using 150-nm diameter Ag nanoparticles. We term the resultant electrodynamically interacting nanoparticles a driven optical matter system. We also show that a superior trap is created by using a Au nanoplate mirror in a retroreflection geometry, which increases the electric field intensity, the optical drive force, and spatial confinement. Using nanoparticles versus micron sized colloids significantly reduces the surface hydrodynamic friction allowing us to access small values of optical topological charge and drive force. We quantify a further 50% reduction of hydrodynamic friction when the nanoparticles are driven over the Au nanoplate mirrors versus over a mildly electrostatically repulsive glass surface. Further, we demonstrate through experiments and electrodynamics–Langevin dynamics simulations that the optical drive force and the interparticle interactions are not constant around the ring for linearly polarized light, resulting in a strong position-dependent variation in the nanoparticle velocity. The nonuniformity in the optical drive force is also manifest as an increase in fluctuations of interparticle separation, or effective temperature, as the optical driving force is increased. Finally, we resolve an open issue in the literature on periodic modulation of interparticle separation with comparative measurements of driven 300-nm-diameter polystyrene beads that also clearly reveal the significance of electrodynamic forces and interactions in optically driven colloidal systems. Therefore, the modulations in the optical forces and electrodynamic interactions that we demonstrate should not be neglected for dielectric particles and might give rise to some structural and dynamic features that have previously been attributed exclusively to hydrodynamic interactions.

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Friday, February 10, 2017

Direct measurement of sequence-dependent transition path times and conformational diffusion in DNA duplex formation

Krishna Neupane, Feng Wang, and Michael T. Woodside

The conformational diffusion coefficient, D, sets the timescale for microscopic structural changes during folding transitions in biomolecules like nucleic acids and proteins. D encodes significant information about the folding dynamics such as the roughness of the energy landscape governing the folding and the level of internal friction in the molecule, but it is challenging to measure. The most sensitive measure of D is the time required to cross the energy barrier that dominates folding kinetics, known as the transition path time. To investigate the sequence dependence of D in DNA duplex formation, we measured individual transition paths from equilibrium folding trajectories of single DNA hairpins held under tension in high-resolution optical tweezers. Studying hairpins with the same helix length but with G:C base-pair content varying from 0 to 100%, we determined both the average time to cross the transition paths, τtp, and the distribution of individual transit times, PTP(t). We then estimated D from both τtp and PTP(t) from theories assuming one-dimensional diffusive motion over a harmonic barrier. τtp decreased roughly linearly with the G:C content of the hairpin helix, being 50% longer for hairpins with only A:T base pairs than for those with only G:C base pairs. Conversely, D increased linearly with helix G:C content, roughly doubling as the G:C content increased from 0 to 100%. These results reveal that G:C base pairs form faster than A:T base pairs because of faster conformational diffusion, possibly reflecting lower torsional barriers, and demonstrate the power of transition path measurements for elucidating the microscopic determinants of folding.

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Scattering Forces within a Left-Handed Photonic Crystal

Angeleene S. Ang, Sergey V. Sukhov, Aristide Dogariu & Alexander S. Shalin

Electromagnetic waves are known to exert optical forces on particles through radiation pressure. It was hypothesized previously that electromagnetic waves inside left-handed metamaterials produce negative radiation pressure. Here we numerically examine optical forces inside left-handed photonic crystals demonstrating negative refraction and reversed phase propagation. We demonstrate that even though the direction of force might not follow the flow of energy, the positive radiation pressure is maintained inside photonic crystals.

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Manipulating orbital angular momentum of light with tailored in-plane polarization states

Luping Du, Zhongsheng Man, Yuquan Zhang, Changjun Min, Siwei Zhu & Xiaocong Yuan

Generally, polarization and phase are considered as two relatively independent parameters of light, and show little interaction when a light propagates in a homogeneous and isotropic medium. Here, we reveal that orbital angular momentum (OAM) of an optical vortex beam can be modulated by specially-tailored locally linear polarization states of light under a tightly-focusing conditon. We perform both theoretical and experimental studies of this interaction between vortex phase and vector polarization, and find that an arbitrary topological charge value of OAM can be achieved in principle through vector polarization modulation, in contrast to the spin-orbital conversion that yields only the ± ћ OAM values through circular polarization. We verify the modulation of optical OAM state with vector beams by observing the orbital rotation of trapped particles.

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Radiation force of a high-energy laser and its effects on second-harmonic generation

Ruifeng Su, Haitao Liu, Yingchun Liang, and Fuli Yu

The radiation force of a high-energy laser caused by reflection at the input surface of a mounted KH2PO4KH2PO4 (KDP) crystal is studied, along with its effects on the second-harmonic generation (SHG) efficiency of the laser beam. A comprehensive model incorporating principles of momentum transfer, mechanics, and optics is proposed, taking advantage of which, the mechanical stress within the KDP crystal that is caused by the radiation force, and the SHG efficiency that is affected by the stress are successively studied. Moreover, the effects of the intensity of the laser beam on the radiation force, the stress, and the SHG efficiency are determined, respectively. It demonstrates that a high-energy laser beam causes macroscopic radiation force and further contributes negative effects to SHG efficiency.

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Cofilin Regulates Nuclear Architecture through a Myosin-II Dependent Mechanotransduction Module

O’Neil Wiggan, Bryce Schroder, Diego Krapf, James R. Bamburg & Jennifer G. DeLuca

Structural features of the nucleus including shape, size and deformability impact its function affecting normal cellular processes such as cell differentiation and pathological conditions such as tumor cell migration. Despite the fact that abnormal nuclear morphology has long been a defining characteristic for diseases such as cancer relatively little is known about the mechanisms that control normal nuclear architecture. Mounting evidence suggests close coupling between F-actin cytoskeletal organization and nuclear morphology however, mechanisms regulating this coupling are lacking. Here we identify that Cofilin/ADF-family F-actin remodeling proteins are essential for normal nuclear structure in different cell types. siRNA mediated silencing of Cofilin/ADF provokes striking nuclear defects including aberrant shapes, nuclear lamina disruption and reductions to peripheral heterochromatin. We provide evidence that these anomalies are primarily due to Rho kinase (ROCK) controlled excessive contractile myosin-II activity and not to elevated F-actin polymerization. Furthermore, we demonstrate a requirement for nuclear envelope LINC (linker of nucleoskeleton and cytoskeleton) complex proteins together with lamin A/C for nuclear aberrations induced by Cofilin/ADF loss. Our study elucidates a pivotal regulatory mechanism responsible for normal nuclear structure and which is expected to fundamentally influence nuclear function.

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