Blog
About

4
views
0
recommends
+1 Recommend
0 collections
    0
    shares
      • Record: found
      • Abstract: found
      • Article: found
      Is Open Access

      SMTracker: a tool for quantitative analysis, exploration and visualization of single-molecule tracking data reveals highly dynamic binding of B. subtilis global repressor AbrB throughout the genome

      Read this article at

      Bookmark
          There is no author summary for this article yet. Authors can add summaries to their articles on ScienceOpen to make them more accessible to a non-specialist audience.

          Abstract

          Single-particle (molecule) tracking (SPT/SMT) is a powerful method to study dynamic processes in living cells at high spatial and temporal resolution. Even though SMT is becoming a widely used method in bacterial cell biology, there is no program employing different analytical tools for the quantitative evaluation of tracking data. We developed SMTracker, a MATLAB-based graphical user interface (GUI) for automatically quantifying, visualizing and managing SMT data via five interactive panels, allowing the user to interactively explore tracking data from several conditions, movies and cells on a track-by-track basis. Diffusion constants are calculated a) by a Gaussian mixture model (GMM) panel, analyzing the distribution of positional displacements in x- and y-direction using a multi-state diffusion model (e.g. DNA-bound vs. freely diffusing molecules), and inferring the diffusion constants and relative fraction of molecules in each state, or b) by square displacement analysis (SQD), using the cumulative probability distribution of square displacements to estimate the diffusion constants and relative fractions of up to three diffusive states, or c) through mean-squared displacement (MSD) analyses, allowing the discrimination between Brownian, sub- or superdiffusive behavior. A spatial distribution analysis (SDA) panel analyzes the subcellular localization of molecules, summarizing the localization of trajectories in 2D- heat maps. Using SMTracker, we show that the global transcriptional repressor AbrB performs highly dynamic binding throughout the Bacillus subtilis genome, with short dwell times that indicate high on/off rates in vivo. While about a third of AbrB molecules are in a DNA-bound state, 40% diffuse through the chromosome, and the remaining molecules freely diffuse through the cells. AbrB also forms one or two regions of high intensity binding on the nucleoids, similar to the global gene silencer H-NS in Escherichia coli, indicating that AbrB may also confer a structural function in genome organization.

          Related collections

          Most cited references 32

          • Record: found
          • Abstract: found
          • Article: not found

          Probing transcription factor dynamics at the single-molecule level in a living cell.

           X. C. Xie,  Wei Li,  Johan Elf (2007)
          Transcription factors regulate gene expression through their binding to DNA. In a living Escherichia coli cell, we directly observed specific binding of a lac repressor, labeled with a fluorescent protein, to a chromosomal lac operator. Using single-molecule detection techniques, we measured the kinetics of binding and dissociation of the repressor in response to metabolic signals. Furthermore, we characterized the nonspecific binding to DNA, one-dimensional (1D) diffusion along DNA segments, and 3D translocation among segments through cytoplasm at the single-molecule level. In searching for the operator, a lac repressor spends approximately 90% of time nonspecifically bound to and diffusing along DNA with a residence time of <5 milliseconds. The methods and findings can be generalized to other nucleic acid binding proteins.
            Bookmark
            • Record: found
            • Abstract: found
            • Article: not found

            High-throughput, subpixel precision analysis of bacterial morphogenesis and intracellular spatio-temporal dynamics.

            Bacteria display various shapes and rely on complex spatial organization of their intracellular components for many cellular processes. This organization changes in response to internal and external cues. Quantitative, unbiased study of these spatio-temporal dynamics requires automated image analysis of large microscopy datasets. We have therefore developed MicrobeTracker, a versatile and high-throughput image analysis program that outlines and segments cells with subpixel precision, even in crowded images and mini-colonies, enabling cell lineage tracking. MicrobeTracker comes with an integrated accessory tool, SpotFinder, which precisely tracks foci of fluorescently labelled molecules inside cells. Using MicrobeTracker, we discover that the dynamics of the extensively studied Escherichia coli Min oscillator depends on Min protein concentration, unveiling critical limitations in robustness within the oscillator. We also find that the fraction of MinD proteins oscillating increases with cell length, indicating that the oscillator has evolved to be most effective when cells attain an appropriate length. MicrobeTracker was also used to uncover novel aspects of morphogenesis and cell cycle regulation in Caulobacter crescentus. By tracking filamentous cells, we show that the chromosomal origin at the old-pole is responsible for most replication/separation events while the others remain largely silent despite contiguous cytoplasm. This surprising position-dependent silencing is regulated by division. © 2011 Blackwell Publishing Ltd.
              Bookmark
              • Record: found
              • Abstract: found
              • Article: found
              Is Open Access

              Objective comparison of particle tracking methods

              The first community competition designed to objectively compare the performance of particle tracking algorithms provides valuable practical information for both users and developers. Supplementary information The online version of this article (doi:10.1038/nmeth.2808) contains supplementary material, which is available to authorized users.
                Bookmark

                Author and article information

                Contributors
                georg.fritz@synmikro.uni-marburg.de
                peter.graumann@synmikro.uni-marburg.de
                Journal
                Sci Rep
                Sci Rep
                Scientific Reports
                Nature Publishing Group UK (London )
                2045-2322
                24 October 2018
                24 October 2018
                2018
                : 8
                Affiliations
                [1 ]GRID grid.452532.7, SYNMIKRO, LOEWE Center for Synthetic Microbiology, ; Marburg, Germany
                [2 ]ISNI 0000 0004 1936 9756, GRID grid.10253.35, Department of Chemistry, , Philipps Universität Marburg, ; Marburg, Germany
                [3 ]ISNI 0000 0004 1936 9756, GRID grid.10253.35, Department of Physics, , Philipps Universität Marburg, ; Marburg, Germany
                Article
                33842
                10.1038/s41598-018-33842-9
                6200787
                30356068
                © The Author(s) 2018

                Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

                Categories
                Article
                Custom metadata
                © The Author(s) 2018

                Uncategorized

                Comments

                Comment on this article