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Single molecule localization

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https://www.readbyqxmd.com/read/28448050/single-molecule-analysis-of-laser-localized-psoralen-adducts
#1
Jing Huang, Himabindu Gali, Julia Gichimu, Marina A Bellani, Durga Pokharel, Manikandan Paramasivam, Michael M Seidman
The DNA Damage Response (DDR) has been extensively characterized in studies of double strand breaks (DSBs) induced by laser micro beam irradiation in live cells. The DDR to helix distorting covalent DNA modifications, including interstrand DNA crosslinks (ICLs), is not as well defined. We have studied the DDR stimulated by ICLs, localized by laser photoactivation of immunotagged psoralens, in the nuclei of live cells. In order to address fundamental questions about adduct distribution and replication fork encounters, we combined laser localization with two other technologies...
April 20, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/28441775/progress-in-the-correlative-atomic-force-microscopy-and-optical-microscopy
#2
REVIEW
Lulu Zhou, Mingjun Cai, Ti Tong, Hongda Wang
Atomic force microscopy (AFM) has evolved from the originally morphological imaging technique to a powerful and multifunctional technique for manipulating and detecting the interactions between molecules at nanometer resolution. However, AFM cannot provide the precise information of synchronized molecular groups and has many shortcomings in the aspects of determining the mechanism of the interactions and the elaborate structure due to the limitations of the technology, itself, such as non-specificity and low imaging speed...
April 24, 2017: Sensors
https://www.readbyqxmd.com/read/28439848/the-janus-head-of-oxidative-stress-in-metabolic-diseases-and-during-physical-exercise
#3
REVIEW
Dominik Pesta, Michael Roden
PURPOSE OF REVIEW: Oxidative stress describes an imbalance between production and degradation of reactive oxygen species (ROS), which can damage macromolecules. However, ROS may also serve as signaling molecules activating cellular pathways involved in cell proliferation and adaptation. This review describes alterations in metabolic diseases including obesity, insulin resistance, and/or diabetes mellitus as well as responses to acute and chronic physical exercise. RECENT FINDINGS: Chronic upregulation of oxidative stress associates with the development of insulin resistance and type 2 diabetes (T2D)...
June 2017: Current Diabetes Reports
https://www.readbyqxmd.com/read/28437460/an-aggregation-removal-model-for-the-formation-and-size-determination-of-post-synaptic-scaffold-domains
#4
Jonas Ranft, Leandro G Almeida, Pamela C Rodriguez, Antoine Triller, Vincent Hakim
The formation and stability of synapses are key questions in neuroscience. Post-synaptic domains have been classically conceived as resulting from local insertion and turnover of proteins at the synapse. However, insertion is likely to occur outside the post-synaptic domains and advances in single-molecule imaging have shown that proteins diffuse in the plane of the membrane prior to their accumulation at synapses. We quantitatively investigated this scenario using computer simulations and mathematical analysis, taking for definiteness the specific case of inhibitory synapse components, i...
April 24, 2017: PLoS Computational Biology
https://www.readbyqxmd.com/read/28437075/fluorescent-photoswitchable-diarylethenes-for-biolabeling-and-single-molecule-localization-microscopies-with-optical-superresolution
#5
Benoit Roubinet, Michael Weber, Heydar Shojaei, Mark Bates, Mariano L Bossi, Vladimir N Belov, Masahiro Irie, Stefan W Hell
A modular assembly of water soluble diarylethenes (DAEs), applicable as bio-markers for optical nanoscopy, is reported. Reversibly photoswitchable 1,2-bis(2-alkyl-6-phenyl-1-benzothiophene-1,1-dioxide-3-yl)perfluorocyclopentenes possessing a fluorescent "closed" form were decorated with one or two methoxy group(s) attached to the para-position(s) of phenyl ring(s) and two, four or eight carboxylic acid groups. Antibody conjugates of these DAEs feature low aggregation, efficient photoswitching in aqueous buffers, specific staining of cellular structures and photophysical properties (high emission efficiencies and low cycloreversion quantum yields) enabling their application in superresolution microscopy...
April 24, 2017: Journal of the American Chemical Society
https://www.readbyqxmd.com/read/28435614/time-resolved-biophysical-approaches-to-nucleocytoplasmic-transport
#6
REVIEW
Francesco Cardarelli
Molecules are continuously shuttling across the nuclear envelope barrier that separates the nucleus from the cytoplasm. Instead of being just a barrier to diffusion, the nuclear envelope is rather a complex filter that provides eukaryotes with an elaborate spatiotemporal regulation of fundamental molecular processes, such as gene expression and protein translation. Given the highly dynamic nature of nucleocytoplasmic transport, during the past few decades large efforts were devoted to the development and application of time resolved, fluorescence-based, biophysical methods to capture the details of molecular motion across the nuclear envelope...
2017: Computational and Structural Biotechnology Journal
https://www.readbyqxmd.com/read/28432305/systematic-synergy-of-glucose-and-glp-1-to-stimulate-insulin-secretion-revealed-by-quantitative-phosphoproteomics
#7
Jia-Shu Tang, Qing-Run Li, Jia-Ming Li, Jia-Rui Wu, Rong Zeng
GLP-1 synergizes with glucose in regulating pancreatic β-cell function, including facilitating β-cell survival and insulin secretion. Though it has been widely accepted that phosphorylation is extremely important in regulating β-cell functions, our knowledge to the global mechanism is still limited. Here we performed a quantitative phosphoproteomics study to systematically present the synergistic regulation of INS-1E cell phosphoproteome mediated by glucose and GLP-1. We generated the largest pancreatic β-cell phosphoproteome by identifying 25,327 accurately localized phosphorylation sites on 5,389 proteins...
April 21, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28431428/computational-and-in-vitro-experimental-investigation-of-intrathecal-drug-distribution-parametric-study-of-the-effect-of-injection-volume-cerebrospinal-fluid-pulsatility-and-drug-uptake
#8
Kevin M Tangen, Roxanne Leval, Ankit I Mehta, Andreas A Linninger
BACKGROUND: Intrathecal drug delivery is an attractive option to circumvent the blood-brain barrier for pain management through its increased efficacy of pain relief, reduction in adverse side effects, and cost-effectiveness. Unfortunately, there are limited guidelines for physicians to choose infusion or drug pump settings to administer therapeutic doses to specific regions of the spine or the brain. Although empiric trialing of intrathecal drugs is critical to determine the sustained side effects, currently there is no inexpensive in vitro method to guide the selection of spinal drug delivery parameters...
May 2017: Anesthesia and Analgesia
https://www.readbyqxmd.com/read/28430483/infrared-peak-splitting-from-phonon-localization-in-solid-hydrogen
#9
Ioan B Magdău, Graeme J Ackland
We show that the isotope effect leads to a completely different spectroscopic signal in hydrogen-deuterium mixtures, compared to pure elements that have the same crystal structure. This is particularly true for molecular vibrations, which are the main source of information about the structure of high-pressure hydrogen. Mass disorder breaks translational symmetry, meaning that vibrations are localized almost to single molecules, and are not zone-center phonons. In mixtures, each observable infrared (IR) peak corresponds to a collection of many such molecular vibrations, which have a distribution of frequencies depending on local environment...
April 7, 2017: Physical Review Letters
https://www.readbyqxmd.com/read/28424492/teststorm-versatile-simulator-software-for-multimodal-super-resolution-localization-fluorescence-microscopy
#10
Tibor Novák, Tamás Gajdos, József Sinkó, Gábor Szabó, Miklós Erdélyi
Optimization of sample, imaging and data processing parameters is an essential task in localization based super-resolution microscopy, where the final image quality strongly depends on the imaging of single isolated fluorescent molecules. A computational solution that uses a simulator software for the generation of test data stacks was proposed, developed and tested. The implemented advanced physical models such as scalar and vector based point spread functions, polarization sensitive detection, drift, spectral crosstalk, structured background etc...
April 19, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28424418/the-aryl-hydrocarbon-receptor-is-required-for-induction-of-p21cip1-waf1-expression-and-growth-inhibition-by-su5416-in-hepatoma-cells
#11
Edmond F O'Donnell, Hyo Sang Jang, Martin Pearce, Nancy I Kerkvliet, Siva Kumar Kolluri
The aryl hydrocarbon receptor (AhR) is a potential clinical target for cancer and autoimmune dysfunction. Identifying selective AhR modulators that produce desirable clinical outcomes represents an opportunity for developing new anti-cancer agents. Repurposing clinically-used drugs with established safety profiles that activate the AhR represents a good starting place to pursue this goal. In this study, we characterized the AhR-dependent effects of SU5416 (Semaxanib) following its identification in a small-molecule library screen...
April 11, 2017: Oncotarget
https://www.readbyqxmd.com/read/28423324/resolving-subcellular-mirna-trafficking-and-turnover-at-single-molecule-resolution
#12
Sethuramasundaram Pitchiaya, Laurie A Heinicke, Jun I Park, Elizabeth L Cameron, Nils G Walter
Regulation of microRNA (miRNA) localization and stability is critical for their extensive cytoplasmic RNA silencing activity and emerging nuclear functions. Here, we have developed single-molecule fluorescence-based tools to assess the subcellular trafficking, integrity, and activity of miRNAs. We find that seed-matched RNA targets protect miRNAs against degradation and enhance their nuclear retention. While target-stabilized, functional, cytoplasmic miRNAs reside in high-molecular-weight complexes, nuclear miRNAs, as well as cytoplasmic miRNAs targeted by complementary anti-miRNAs, are sequestered stably within significantly lower-molecular-weight complexes and rendered repression incompetent...
April 18, 2017: Cell Reports
https://www.readbyqxmd.com/read/28404604/unusual-semi-extractability-as-a-hallmark-of-nuclear-body-associated-architectural-noncoding-rnas
#13
Takeshi Chujo, Tomohiro Yamazaki, Tetsuya Kawaguchi, Satoshi Kurosaka, Toru Takumi, Shinichi Nakagawa, Tetsuro Hirose
NEAT1_2 long noncoding RNA (lncRNA) is the molecular scaffold of paraspeckle nuclear bodies. Here, we report an improved RNA extraction method: extensive needle shearing or heating of cell lysate in RNA extraction reagent improved NEAT1_2 extraction by 20-fold (a property we term "semi-extractability"), whereas using a conventional method NEAT1_2 was trapped in the protein phase. The improved extraction method enabled us to estimate that approximately 50 NEAT1_2 molecules are present in a single paraspeckle...
April 12, 2017: EMBO Journal
https://www.readbyqxmd.com/read/28404480/2d-and-3d-fish-of-expanded-repeat-rnas-in-human-lymphoblasts
#14
Martyna O Urbanek, Michał Michalak, Wlodzimierz J Krzyzosiak
The first methods for visualizing RNAs within cells were designed for simple imaging of specific transcripts in cells or tissues and since then significant technical advances have been made in this field. Today, high-resolution images can be obtained, enabling visualization of single transcript molecules, quantitative analyses of images, and precise localization of RNAs within cells as well as co-localization of transcripts with specific proteins or other molecules. In addition, tracking of RNA dynamics within single cell has become possible...
April 9, 2017: Methods: a Companion to Methods in Enzymology
https://www.readbyqxmd.com/read/28402886/three-dimensional-super-resolution-in-eukaryotic-cells-using-the-double-helix-point-spread-function
#15
Alexander R Carr, Aleks Ponjavic, Srinjan Basu, James McColl, Ana Mafalda Santos, Simon Davis, Ernest D Laue, David Klenerman, Steven F Lee
Single-molecule localization microscopy, typically based on total internal reflection illumination, has taken our understanding of protein organization and dynamics in cells beyond the diffraction limit. However, biological systems exist in a complicated three-dimensional environment, which has required the development of new techniques, including the double-helix point spread function (DHPSF), to accurately visualize biological processes. The application of the DHPSF approach has so far been limited to the study of relatively small prokaryotic cells...
April 11, 2017: Biophysical Journal
https://www.readbyqxmd.com/read/28402875/tracking-low-copy-transcription-factors-in-living-bacteria-the-case-of-the-lac-repressor
#16
Federico Garza de Leon, Laura Sellars, Mathew Stracy, Stephen J W Busby, Achillefs N Kapanidis
Transcription factors control the expression of genes by binding to specific sites in DNA and repressing or activating transcription in response to stimuli. The lac repressor (LacI) is a well characterized transcription factor that regulates the ability of bacterial cells to uptake and metabolize lactose. Here, we study the intracellular mobility and spatial distribution of LacI in live bacteria using photoactivated localization microscopy combined with single-particle tracking. Since we track single LacI molecules in live cells by stochastically photoactivating and observing fluorescent proteins individually, there are no limitations on the copy number of the protein under study; as a result, we were able to study the behavior of LacI in bacterial strains containing the natural copy numbers (∼40 monomers), as well as in strains with much higher copy numbers due to LacI overexpression...
April 11, 2017: Biophysical Journal
https://www.readbyqxmd.com/read/28395669/single-swim-sessions-in-c-elegans-induce-key-features-of-mammalian-exercise
#17
Ricardo Laranjeiro, Girish Harinath, Daniel Burke, Bart P Braeckman, Monica Driscoll
BACKGROUND: Exercise exerts remarkably powerful effects on metabolism and health, with anti-disease and anti-aging outcomes. Pharmacological manipulation of exercise benefit circuits might improve the health of the sedentary and the aging populations. Still, how exercised muscle signals to induce system-wide health improvement remains poorly understood. With a long-term interest in interventions that promote animal-wide health improvement, we sought to define exercise options for Caenorhabditis elegans...
April 10, 2017: BMC Biology
https://www.readbyqxmd.com/read/28391361/ultrastructural-localization-of-5-methylcytosine-on-dna-and-rna
#18
Irene Masiello, Marco Biggiogera
DNA methylation is the major epigenetic modification and it is involved in the negative regulation of gene expression. Its alteration can lead to neoplastic transformation. Several biomolecular approaches are nowadays used to study this modification on DNA, but also on RNA molecules, which are known to play a role in different biological processes. RNA methylation is one of the most common RNA modifications and 5-methylcytosine presence has recently been suggested in mRNA. However, an analysis of nucleic acid methylation at electron microscope is still lacking...
April 8, 2017: Cellular and Molecular Life Sciences: CMLS
https://www.readbyqxmd.com/read/28390838/super-resolution-microscopy-approaches-to-nuclear-nanostructure-imaging
#19
Christoph Cremer, Aleksander Szczurek, Florian Schock, Amine Gourram, Udo Birk
The human genome has been decoded, but we are still far from understanding the regulation of all gene activities. A largely unexplained role in these regulatory mechanisms is played by the spatial organization of the genome in the cell nucleus which has far-reaching functional consequences for gene regulation. Until recently, it appeared to be impossible to study this problem on the nanoscale by light microscopy. However, novel developments in optical imaging technology have radically surpassed the limited resolution of conventional far-field fluorescence microscopy (ca...
April 5, 2017: Methods: a Companion to Methods in Enzymology
https://www.readbyqxmd.com/read/28390361/the-role-of-li-ions-in-the-gas-phase-dehydrohalogenation-and-dehydration-reactions-of-i-c3h7br-and-i-c3h7oh-molecules-studied-by-radiofrequency-guided-ion-beam-techniques-and-ab-initio-methods
#20
E López, J M Lucas, J de Andrés, M Albertí, J M Bofill, A Aguilar
Gas phase reactive collisions between lithium ions and i-C3H7X (X = Br, OH) molecules have been studied under single collision conditions in the center of mass (CM) 0.01-10.00 eV energy range using a radiofrequency-guided ion beam apparatus. Mass spectrometry analysis of the products did show the presence of [C3H6-Li](+), [HX-Li](+), C3H7(+), and C2H3(+) as well as of the [Li-i-C3H7Br](+) adduct while [Li-i-C3H7OH](+) was hardly detected. For all these reactive processes, the corresponding cross sections have been measured in absolute units as a function of the CM collision energy...
April 7, 2017: Journal of Chemical Physics
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