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Eileen F Breslin, Jessica Cornell, Zachary Schuhmacher, Madison Himelright, Aya Andos, Ariel Childs, Ana Clem, Monica Gerber, Arissa Gordillo, Laith Harb, Reafa Hossain, Taylor Hutchinson, Isaac Miller, Edmund Morton, Ryan Walters, Destin Webb, Louise Temple
Soil from George Town, Grand Cayman Island, yielded the bacteriophage Belinda, isolated on Bacillus thuringiensis DSM 350. We present here the analysis of the complete genome sequence of 162,308 bp, with 298 predicted genes. The genome also contains three tRNA genes. Belinda belongs to the C1 cluster of Bacillus phages.
October 13, 2016: Genome Announcements
Julie O'Neal, Amy Clem, Lindsey Reynolds, Susan Dougherty, Yoannis Imbert-Fernandez, Sucheta Telang, Jason Chesney, Brian F Clem
PURPOSE: Human epidermal growth factor receptor-2 (HER2) has been implicated in the progression of multiple tumor types, including breast cancer, and many downstream effectors of HER2 signaling are primary regulators of cellular metabolism, including Ras and Akt. A key downstream metabolic target of Ras and Akt is the 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 isozyme (PFKFB3), whose product, fructose-2,6-bisphosphate (F26BP), is a potent allosteric activator of a rate-limiting enzyme in glycolysis, 6-phosphofructo-1-kinase (PFK-1)...
November 2016: Breast Cancer Research and Treatment
Daniel O Daley, Ulf Skoglund, Bill Söderström
The source of constriction required for division of a bacterial cell remains enigmatic. FtsZ is widely believed to be a key player, because in vitro experiments indicate that it can deform liposomes when membrane tethered. However in vivo evidence for such a role has remained elusive as it has been challenging to distinguish the contribution of FtsZ from that of peptidoglycan-ingrowth. To differentiate between these two possibilities we studied the early stages of division in Escherichia coli, when FtsZ is present at the division site but peptidoglycan synthesizing enzymes such as FtsI and FtsN are not...
2016: Scientific Reports
Isabell Begemann, Milos Galic
Many core paradigms of contemporary neuroscience are based on information obtained by electron or light microscopy. Intriguingly, these two imaging techniques are often viewed as complementary, yet separate entities. Recent technological advancements in microscopy techniques, labeling tools, and fixation or preparation procedures have fueled the development of a series of hybrid approaches that allow correlating functional fluorescence microscopy data and ultrastructural information from electron micrographs from a singular biological event...
2016: Frontiers in Synaptic Neuroscience
Christophe Normand, Maxime Berthaud, Olivier Gadal, Isabelle Léger-Silvestre
Nucleoli form around RNA polymerase I transcribed ribosomal RNA (rRNA) genes. The direct electron microscopy observation of rRNA genes after nucleolar chromatin spreading (Miller's spreads) constitutes to date the only system to quantitatively assess transcription at a single molecule level. However, the spreading procedure is likely generating artifact and despite being informative, these spread rRNA genes are far from their in vivo situation. The integration of the structural characterization of spread rRNA genes in the three-dimensional (3D) organization of the nucleolus would represent an important scientific achievement...
2016: Methods in Molecular Biology
P W Gorham, J Nam, A Romero-Wolf, S Hoover, P Allison, O Banerjee, J J Beatty, K Belov, D Z Besson, W R Binns, V Bugaev, P Cao, C Chen, P Chen, J M Clem, A Connolly, B Dailey, C Deaconu, L Cremonesi, P F Dowkontt, M A DuVernois, R C Field, B D Fox, D Goldstein, J Gordon, C Hast, C L Hebert, B Hill, K Hughes, R Hupe, M H Israel, A Javaid, J Kowalski, J Lam, J G Learned, K M Liewer, T C Liu, J T Link, E Lusczek, S Matsuno, B C Mercurio, C Miki, P Miočinović, M Mottram, K Mulrey, C J Naudet, J Ng, R J Nichol, K Palladino, B F Rauch, K Reil, J Roberts, M Rosen, B Rotter, J Russell, L Ruckman, D Saltzberg, D Seckel, H Schoorlemmer, S Stafford, J Stockham, M Stockham, B Strutt, K Tatem, G S Varner, A G Vieregg, D Walz, S A Wissel, F Wu
We report on four radio-detected cosmic-ray (CR) or CR-like events observed with the Antarctic Impulsive Transient Antenna (ANITA), a NASA-sponsored long-duration balloon payload. Two of the four were previously identified as stratospheric CR air showers during the ANITA-I flight. A third stratospheric CR was detected during the ANITA-II flight. Here, we report on characteristics of these three unusual CR events, which develop nearly horizontally, 20-30 km above the surface of Earth. In addition, we report on a fourth steeply upward-pointing ANITA-I CR-like radio event which has characteristics consistent with a primary that emerged from the surface of the ice...
August 12, 2016: Physical Review Letters
Michael R Kanost, Estela L Arrese, Xiaolong Cao, Yun-Ru Chen, Sanjay Chellapilla, Marian R Goldsmith, Ewald Grosse-Wilde, David G Heckel, Nicolae Herndon, Haobo Jiang, Alexie Papanicolaou, Jiaxin Qu, Jose L Soulages, Heiko Vogel, James Walters, Robert M Waterhouse, Seung-Joon Ahn, Francisca C Almeida, Chunju An, Peshtewani Aqrawi, Anne Bretschneider, William B Bryant, Sascha Bucks, Hsu Chao, Germain Chevignon, Jayne M Christen, David F Clarke, Neal T Dittmer, Laura C F Ferguson, Spyridoula Garavelou, Karl H J Gordon, Ramesh T Gunaratna, Yi Han, Frank Hauser, Yan He, Hanna Heidel-Fischer, Ariana Hirsh, Yingxia Hu, Hongbo Jiang, Divya Kalra, Christian Klinner, Christopher König, Christie Kovar, Ashley R Kroll, Suyog S Kuwar, Sandy L Lee, Rüdiger Lehman, Kai Li, Zhaofei Li, Hanquan Liang, Shanna Lovelace, Zhiqiang Lu, Jennifer H Mansfield, Kyle J McCulloch, Tittu Mathew, Brian Morton, Donna M Muzny, David Neunemann, Fiona Ongeri, Yannick Pauchet, Ling-Ling Pu, Ioannis Pyrousis, Xiang-Jun Rao, Amanda Redding, Charles Roesel, Alejandro Sanchez-Gracia, Sarah Schaack, Aditi Shukla, Guillaume Tetreau, Yang Wang, Guang-Hua Xiong, Walther Traut, Tom K Walsh, Kim C Worley, Di Wu, Wenbi Wu, Yuan-Qing Wu, Xiufeng Zhang, Zhen Zou, Hannah Zucker, Adriana D Briscoe, Thorsten Burmester, Rollie J Clem, René Feyereisen, Cornelis J P Grimmelikhuijzen, Stavros J Hamodrakas, Bill S Hansson, Elisabeth Huguet, Lars S Jermiin, Que Lan, Herman K Lehman, Marce Lorenzen, Hans Merzendorfer, Ioannis Michalopoulos, David B Morton, Subbaratnam Muthukrishnan, John G Oakeshott, Will Palmer, Yoonseong Park, A Lorena Passarelli, Julio Rozas, Lawrence M Schwartz, Wendy Smith, Agnes Southgate, Andreas Vilcinskas, Richard Vogt, Ping Wang, John Werren, Xiao-Qiang Yu, Jing-Jiang Zhou, Susan J Brown, Steven E Scherer, Stephen Richards, Gary W Blissard
Manduca sexta, known as the tobacco hornworm or Carolina sphinx moth, is a lepidopteran insect that is used extensively as a model system for research in insect biochemistry, physiology, neurobiology, development, and immunity. One important benefit of this species as an experimental model is its extremely large size, reaching more than 10 g in the larval stage. M. sexta larvae feed on solanaceous plants and thus must tolerate a substantial challenge from plant allelochemicals, including nicotine. We report the sequence and annotation of the M...
September 2016: Insect Biochemistry and Molecular Biology
Matthia A Karreman, Vincent Hyenne, Yannick Schwab, Jacky G Goetz
Studying key biological events within complex model systems relies on dynamic and functional imaging at optimum spatial and temporal resolutions. Intravital correlative light and electron microscopy (intravital CLEM) combines imaging living multicellular model systems with electron microscopy, and offers full ultrastructural details of dynamic or transient events in vivo. However, routine use of intravital CLEM is hindered by multiple technological challenges faced when targeting a micron-size object (e.g., single cells or organelles) in a complex living organism...
August 8, 2016: Trends in Cell Biology
Frank P Macaluso, Geoffrey S Perumal, Johan Kolstrup, Peter Satir
CLEM (correlated light and electron microscope) imaging is a highly useful technique for examining primary cilia. With CLEM, it is possible to determine the distribution of tagged proteins along the ciliary membrane and axoneme with high precision. Scanning electron microscopy (SEM) permits measurement of ciliary length and orientation in relation to nearby cellular structures in a 3D image; in optimal cases, this can be combined with superresolution microscopy of selected ciliary components as they enter or leave the cilium...
2016: Methods in Molecular Biology
Matthew R G Russell, Thomas R Lerner, Jemima J Burden, David O Nkwe, Annegret Pelchen-Matthews, Marie-Charlotte Domart, Joanne Durgan, Anne Weston, Martin L Jones, Christopher J Peddie, Raffaella Carzaniga, Oliver Florey, Mark Marsh, Maximiliano G Gutierrez, Lucy M Collinson
The processes of life take place in multiple dimensions, but imaging these processes in even three dimensions is challenging. Here we describe a workflow for 3D correlative light and electron microscopy (CLEM) of cell monolayers using fluorescence microscopy to identify and follow biological events, combined with serial blockface scanning electron microscopy to analyse the underlying ultrastructure. The workflow encompasses all steps from cell culture to sample processing, imaging strategy and 3D image processing/analysis...
July 21, 2016: Journal of Cell Science
Ekaterina A Souslova, Kristina E Mironova, Sergey M Deyev
Genetically encoded photosensitizers (PSs), e.g. ROS generating proteins, correspond to a novel class of PSs that are highly desirable for biological and medical applications since they can be used in combination with a variety of genetic engineering manipulations allowing for precise spatio-temporal control of ROS production within living cells and organisms. In contrast to the commonly used chemical PSs, they can be modified using genetic engineering approaches and targeted to particular cellular compartments and cell types...
July 20, 2016: Journal of Biophotonics
G Casanova, F Nolin, L Wortham, D Ploton, V Banchet, J Michel
Freeze-drying of cryosections of cells or tissues is considered to be the most efficient preparation for microanalysis purpose related to transmission electron microscopy. It allows the measurements of ions and water contents at the ultrastructural level. However an important drawback is associated to freeze-drying: the shrinkage of the cryosections. The aim of this paper is the investigation of this phenomenon by means of three different methods applied to both hydrated and dehydrated cryosections: direct distance measurements on fiducial points, thickness measurements by energy filtered transmission microscopy (EFTEM) and cryo-correlative light electron microscopy (cryo-CLEM)...
September 2016: Micron: the International Research and Review Journal for Microscopy
Elizabeth K Lucas, Anita M Jegarl, Hirofumi Morishita, Roger L Clem
Stimulus processing in fear conditioning is constrained by parvalbumin interneurons (PV-INs) through inhibition of principal excitatory neurons. However, the contributions of PV-IN microcircuits to input gating and long-term plasticity in the fear system remain unknown. Here we interrogate synaptic connections between afferent pathways, PV-INs, and principal excitatory neurons in the basolateral amygdala. We find that subnuclei of this region are populated two functionally distinct PV-IN networks. PV-INs in the lateral (LA), but not the basal (BA), amygdala possess complex dendritic arborizations, receive potent excitatory drive, and mediate feedforward inhibition onto principal neurons...
August 3, 2016: Neuron
H M van der Schaar, C E Melia, J A C van Bruggen, J R P M Strating, M E D van Geenen, A J Koster, M Bárcena, F J M van Kuppeveld
Like all other positive-strand RNA viruses, enteroviruses generate new organelles (replication organelles [ROs]) with a unique protein and lipid composition on which they multiply their viral genome. Suitable tools for live-cell imaging of enterovirus ROs are currently unavailable, as recombinant enteroviruses that carry genes that encode RO-anchored viral proteins tagged with fluorescent reporters have not been reported thus far. To overcome this limitation, we used a split green fluorescent protein (split-GFP) system, comprising a large fragment [strands 1 to 10; GFP(S1-10)] and a small fragment [strand 11; GFP(S11)] of only 16 residues...
July 2016: MSphere
Shouhei Kobayashi, Masaaki Iwamoto, Tokuko Haraguchi
Fluorescence microscopy (FM) is a powerful tool for observing specific molecular components in living cells, but its spatial resolution is relatively low. In contrast, electron microscopy (EM) provides high-resolution information about cellular structures, but it cannot provide temporal information in living cells. To achieve molecular selectivity in imaging at high resolution, a method combining EM imaging with live-cell fluorescence imaging, known as live correlative light-EM (CLEM), has been developed. In this method, living cells are first observed by FM, fixed in situ during the live observation and then subjected to EM observation...
August 2016: Microscopy
Chunfeng Wu, Zihao Deng, Zhao Long, Yi Cai, Zhongfu Ying, Hanqi Yin, Meijin Yuan, Rollie J Clem, Kai Yang, Yi Pang
As baculoviruses usually have a narrow insecticidal spectrum, knowing the mechanisms by which they control the host-range is prerequisite for improvement of their applications as pesticides. In this study, from supernatant of culture cells transfected with DNAs of an Autographa californica multiple nucleopolyhedrovirus (AcMNPV) mutant lacking the antiapoptotic gene p35 (vAc(∆P35)) and a cosmid representing a fragment of Spodoptera exigua nucleopolyhedrovirus (SeMNPV), a viral strain was plaque-purified and named vAcRev...
2016: Scientific Reports
Alberto Rodríguez-Pulido, Aitziber L Cortajarena, Joaquim Torra, Rubén Ruiz-González, Santi Nonell, Cristina Flors
Photosensitizing flavoproteins have great potential as tags for correlative light and electron microscopy (CLEM). We examine the photostability of miniSOG mutants and their ability to photo-oxidize diaminobenzidine, both key aspects for CLEM. Our experiments reveal a complex relation between these parameters and the production of different reactive oxygen species.
June 28, 2016: Chemical Communications: Chem Comm
Jeroen Kuipers, Ruby D Kalicharan, Anouk H G Wolters, Tjakko J van Ham, Ben N G Giepmans
Large-scale 2D electron microscopy (EM), or nanotomy, is the tissue-wide application of nanoscale resolution electron microscopy. Others and we previously applied large scale EM to human skin pancreatic islets, tissue culture and whole zebrafish larvae(1-7). Here we describe a universally applicable method for tissue-scale scanning EM for unbiased detection of sub-cellular and molecular features. Nanotomy was applied to investigate the healthy and a neurodegenerative zebrafish brain. Our method is based on standardized EM sample preparation protocols: Fixation with glutaraldehyde and osmium, followed by epoxy-resin embedding, ultrathin sectioning and mounting of ultrathin-sections on one-hole grids, followed by post staining with uranyl and lead...
2016: Journal of Visualized Experiments: JoVE
Cassandra L Bolar, Natalie Hernandez, Tabia Henry Akintobi, Calvin McAllister, Aneeqah S Ferguson, Latrice Rollins, Glenda Wrenn, Martha Okafor, David Collins, Thomas Clem
BACKGROUND: Among children, there are substantial ethno-racial minority disparities across a broad range of health-related behaviors, experiences, and outcomes. Addressing these disparities is important, as childhood and adolescence establish health trajectories that extend throughout life. METHODS: The current study employed a community-based participatory research approach to gain community insight on child health priorities and to frame an intervention aimed at improving the health of minority children...
2016: Journal of the Georgia Public Health Association
Daniel M P Ardisson-Araújo, Rayane Nunes Lima, Fernando L Melo, Rollie J Clem, Ning Huang, Sônia Nair Báo, Daniel R Sosa-Gómez, Bergmann M Ribeiro
The genome of a novel group II alphabaculovirus, Perigonia lusca single nucleopolyhedrovirus (PeluSNPV), was sequenced and shown to contain 132,831 bp with 145 putative ORFs (open reading frames) of at least 50 amino acids. An interesting feature of this novel genome was the presence of a putative nucleotide metabolism enzyme-encoding gene (pelu112). The pelu112 gene was predicted to encode a fusion of thymidylate kinase (tmk) and dUTP diphosphatase (dut). Phylogenetic analysis indicated that baculoviruses have independently acquired tmk and dut several times during their evolution...
2016: Scientific Reports
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