Oleg Krichevsky - Academia.edu (original) (raw)

Papers by Oleg Krichevsky

We adapt Fluorescence Correlation spectroscopy (FCS) formalism to the studies of the dynamics of ... more We adapt Fluorescence Correlation spectroscopy (FCS) formalism to the studies of the dynamics of semi-flexible polymers and derive expressions relating FCS correlation function to the longitudinal and transverse mean square displacements of polymer segments. We use the derived expressions to measure the dynamics of actin filaments in two experimental situations: filaments labeled at distinct positions and homogeneously labeled filaments. Both approaches give consistent results and allow to measure the temporal dependence of the segmental mean-square displacement (MSD) over almost five decades in time, from ~0.04ms to 2s. These noninvasive measurements allow for a detailed quantitative comparison of the experimental data to the current theories of semi-flexible polymer dynamics. Good quantitative agreement is found between the experimental results and theories explicitly accounting for the hydrodynamic interactions between polymer segments.

We study the fluctuation dynamics of localized denaturation bubbles in heteropolymer DNA with a m... more We study the fluctuation dynamics of localized denaturation bubbles in heteropolymer DNA with a master equation and complementary stochastic simulation based on novel DNA stability data. A significant dependence of opening probability and waiting time between bubble events on the local DNA sequence is revealed and quantified for a biological sequence of the T7 bacteriophage. Quantitative agreement with data from fluorescence correlation spectroscopy (FCS) is demonstrated.

While the statistical mechanical description of DNA has a long tradition, renewed interest in DNA... more While the statistical mechanical description of DNA has a long tradition, renewed interest in DNA melting from a physics perspective is nourished by measurements of the fluctuation dynamics of local denaturation bubbles by single molecule spectroscopy. The dynamical opening of DNA bubbles (DNA breathing) is supposedly crucial for biological functioning during, for instance, transcription initiation and DNA's interaction with selectively single-stranded DNA binding proteins. Motivated by this, we consider the bubble breathing dynamics in a heteropolymer DNA based on a (2+1)-variable master equation and complementary stochastic Gillespie simulations, providing the bubble size and the position of the bubble along the sequence as a function of time. We utilize new experimental data that independently obtain stacking and hydrogen bonding contributions to DNA stability. We calculate the spectrum of relaxation times and the experimentally measurable autocorrelation function of a fluoro...

Cell Stem Cell, 2020

Adult mammalian hematopoietic stem cells (HSCs) reside in the bone marrow (BM) but can be mobiliz... more Adult mammalian hematopoietic stem cells (HSCs) reside in the bone marrow (BM) but can be mobilized into blood for use in transplantation. HSCs interact with BM niche cells that produce growth factor c-Kit ligand (Kitl/SCF) and chemokine CXCL12, and were thought to be static and sessile. We used two-photon laser scanning microscopy to visualize genetically labeled HSCs in the BM of live mice for several hours. The majority of HSCs showed a dynamic non-spherical morphology and significant motility, undergoing slow processive motion interrupted by short stretches of confined motion. HSCs moved in the perivascular space and showed intermittent close contacts with SCF-expressing perivascular stromal cells. In contrast, mobilization-inducing blockade of CXCL12 receptor CXCR4 and integrins rapidly abrogated HSC motility and shape dynamics in real time. Our results reveal an unexpectedly dynamic nature of HSC residence in the BM and interaction with the SCF+ stromal niche, which is disrupted during HSC mobilization.

Current biology : CB, Jan 9, 2017

Actin turnover is the central driving force underlying lamellipodial motility. The molecular comp... more Actin turnover is the central driving force underlying lamellipodial motility. The molecular components involved are largely known, and their properties have been studied extensively in vitro. However, a comprehensive picture of actin turnover in vivo is still missing. We focus on fragments from fish epithelial keratocytes, which are essentially stand-alone motile lamellipodia. The geometric simplicity of the fragments and the absence of additional actin structures allow us to characterize the spatiotemporal lamellipodial actin organization with unprecedented detail. We use fluorescence recovery after photobleaching, fluorescence correlation spectroscopy, and extraction experiments to show that about two-thirds of the lamellipodial actin diffuses in the cytoplasm with nearly uniform density, whereas the rest forms the treadmilling polymer network. Roughly a quarter of the diffusible actin pool is in filamentous form as diffusing oligomers, indicating that severing and debranching ar...

Soft Matter, 2015

Scanning fluorescence correlation spectroscopy in combination with specific fluorescence labeling... more Scanning fluorescence correlation spectroscopy in combination with specific fluorescence labeling is used to measure different static and dynamic properties of a soft matter system.

Physical Review Letters, 1994

We present results of an experimental study of surfactant-polymer interactions in free-standing s... more We present results of an experimental study of surfactant-polymer interactions in free-standing soap films. The relaxation of thermally induced corrugations of the surfactant monolayers comprising a film is studied by reflectivity and light-scattering methods. Our data show that polymers influence the thickness of the equilibrium state of a soap film and lead to the creation of new metastable states of larger thickness. The overall behavior is consistent with a picture in which chains form depletion layers near the surfactant walls, inducing an attractive interaction between the monolayers at large polymer concentrations.

Physical Review B, 1992

We have studied experimentally the coarsening of two-dimensional soap froths in the presence of p... more We have studied experimentally the coarsening of two-dimensional soap froths in the presence of pinning centers. When the average bubble size is smaller than the average interpin distance, the growth is unaffected. When both dimensions become comparable the froth enters a crossover regime followed by a pinned state where growth stops. The nature of the long-time configurations depends strongly on the size of the pins relative to the size of the plateau borders. For thick pins, the final configurations consist of combinations of Steiner trees of small numbers of points with pins located at vertices of the film network. For thin pins the final average bubble size is larger than in the case of thick pins and the final configurations are much more varied. Qualitatively similar behavior is observed in grain growth in metals with impurities.

Physical Review E, 1997

We study by light scattering methods the behavior of soap films drawn from solutions of different... more We study by light scattering methods the behavior of soap films drawn from solutions of different surfactant concentration. For concentrations just above the critical micelle concentration, micelles induce a depletion attraction and thus a reduction in equilibrium film thickness. For larger surfactant concentrations we observe stepwise thinning, due to the organization of micelles in layers parallel to the film plane.

Physical Review E, 1995

We report results from an experimental study of coarsening in thin layers of succinonitrile in th... more We report results from an experimental study of coarsening in thin layers of succinonitrile in the presence of impurities. By quenching the latter from the liquid phase to different temperatures within the liquid-solid coexistence region, different solid area fractions phi are obtained. As phi is incresed from 0.13 to 0.40, liquidlike order develops among the coarsening crystals due to diffusional

Physical Review E, 1993

Evolving random cellular structures are observed to reach a universal scaling regime. A mean-fiel... more Evolving random cellular structures are observed to reach a universal scaling regime. A mean-field approach to finding fixed-point distributions in cell-side number is extended to distributions for the average area of cells with a given number of sides. This approach leads to simplified equations that can be analyzed analytically and numerically. The theory's results are compared to experimental results on dynamics and distributions in soap froths and good agreement is achieved.

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Physical Review Letters, 2007

Frontiers in immunology, 2012

In this review we discuss how the competition for cytokines between different cells of the immune... more In this review we discuss how the competition for cytokines between different cells of the immune system can shape the system wide immune response. We focus on interleukin-2 (IL-2) secretion by activated effector T cells (T(eff)) and on the competition for IL-2 consumption between T(eff) and regulatory T cells (T(reg)). We discuss the evidence for the mechanism in which the depletion of IL-2 by T(reg) cells would be sufficient to suppress an autoimmune response, yet not strong enough to prevent an immune response. We present quantitative estimations and summarize our modeling effort to show that the tug-of-war between T(reg) and T(eff) cells for IL-2 molecules can be won by T(reg) cells in the case of weak activation of T(eff) leading to the suppression of the immune response. Or, for strongly activated T(eff) cells, it can be won by T(eff) cells bringing about the activation of the whole adaptive immune system. Finally, we discuss some recent applications attempting to achieve clin...

MRS Proceedings, 2005

Optical tweezers, atomic force microscopes, patch clamping, or fluorescence techniques make it po... more Optical tweezers, atomic force microscopes, patch clamping, or fluorescence techniques make it possible to study both the equilibrium conformations and dynamics of single DNA molecules as well as their interaction with binding proteins. In this paper we address the dynamics of local DNA denaturation (bubble breathing), deriving its dynamic response to external physical parameters and the DNA sequence in terms of the bubble relaxation time spectrum and the autocorrelation function of bubble breathing. The interaction with binding proteins that selectively bind to the DNA single strand exposed in a denaturation bubble are shown to involve an interesting competition of time scales, varying between kinetic blocking of protein binding up to full binding protein-induced denaturation of the DNA. We will also address the potential to use DNA physics for the design of nanosensors. Finally, we report recent findings on the search process of proteins for a specific target on the DNA.

eLife, 2014

Variability within isogenic T cell populations yields heterogeneous ‘local’ signaling responses t... more Variability within isogenic T cell populations yields heterogeneous ‘local’ signaling responses to shared antigenic stimuli, but responding clones may communicate ‘global’ antigen load through paracrine messengers, such as cytokines. Such coordination of individual cell responses within multicellular populations is critical for accurate collective reactions to shared environmental cues. However, cytokine production may saturate as a function of antigen input, or be dominated by the precursor frequency of antigen-specific T cells. Surprisingly, we found that T cells scale their collective output of IL-2 to total antigen input over a large dynamic range, independently of population size. Through experimental quantitation and computational modeling, we demonstrate that this scaling is enforced by an inhibitory cross-talk between antigen and IL-2 signaling, and a nonlinear acceleration of IL-2 secretion per cell. Our study reveals how time-integration of these regulatory loops within in...

Using a new method we measure the structure of solutions of semi-exible polymers. In our approach... more Using a new method we measure the structure of solutions of semi-exible polymers. In our approach the sample is uorescently labeled, and uctuations in uorescence are monitored as the sample is scanned through a confocal volume at high speed. We show that in such conditions, the temporal correlation of uorescence uctuations reects the spatial correlations in polymer organization, characterized by the static structure factor. We apply our method to studies of DNA solutions, both in dilute regime, where polymer coils are well separated, and in semi-dilute regime, where polymer coils interpenetrate. In the dilute regime our data is in the excellent agreement with the results of our simulations of DNA structure with known DNA parameters. In semi-dilute regime, we observe the screening eect of polymer mesh on the solutions structure. As concentration in the solution increases, the smaller the screening length is. The dependence of the screening length on DNA concentration follows the pred...

Reports on Progress in Physics, 2002

Fluorescence correlation spectroscopy (FCS) is an experimental technique using statistical analys... more Fluorescence correlation spectroscopy (FCS) is an experimental technique using statistical analysis of the fluctuations of fluorescence in a system in order to decipher dynamic molecular events, such as diffusion or conformational fluctuations of biomolecules. First introduced by Magde et al to measure the diffusion and binding of ethidium bromide onto double-stranded DNA, the technique has been undergoing a renaissance since 1993 with the implementation of confocal microscopy FCS. Since then, a flurry of experiments has implemented FCS to characterize the photochemistry of dyes, the translational and rotational mobilities of fluorescent molecules, as well as to monitor conformational fluctuations of green fluorescent proteins and DNA molecules. In this review, we present the analytical formalism of an FCS measurement, as well as practical considerations for the design of an FCS setup and experiment. We then review the recent applications of FCS in analytical chemistry, biophysics and cell biology, specifically emphasizing the advantages and pitfalls of the technique compared to alternative spectroscopic tools. We also discuss recent extensions of FCS in single-molecule spectroscopy, offering alternative data processing of fluorescence signals to glean more information on the kinetic processes.