Governing fungal polar cell extension: Analysis of Rho GTPase and NDR kinase signalling in Neurospora crassa

Die Entwicklung gestreckter Zelltypen wie Nervenzellen, Pollenschläuche und Zellen filamentöser Pilze erfordert polare Gestaltbildung. Die grundlegenden Signalwege dafür sind hochkonserviert. Missbildungen der Zellform können zu Entwicklungsstörungen oder dem Tod der betroffenen Zelle führen. In dieser Arbeit werden Komponenten, die für polares Wachstum von Neurospora crassa benötigt werden, auf molekularer Ebene charakterisiert. Dabei handelt es sich zum einen um ein Mitglied der GTPase aktivierenden Proteine, LRG1 und zum anderen um die "germinal center" Kinase POD6.POD6 und LRG1 sind für das gerichtete Hyphenwachstum unabdingbar. Deletionsund temperatur-sensitive Allele der Gene cot-1, pod-6 und lrg-1 zeigen phänotypisch Ähnlichkeiten bezüglich des Abbruchs polaren Wachstums, übermäßiger Verzweigungsbildung und der veränderten Größe der Hyphenkompartemente. Komplementationsexperimente belegen, das sowohl der die drei LIM Domänen enthaltende N-Terminus als auch der die Rho-GAP Domäne enthaltende C-Terminus von LRG1 für dessen Funktion benötigt werden. Neben genetischen Hinweisen identifizieren in-vitro Untersuchungen LRG1 als Rho1 spezifisches GAP.COT1, POD6 und LRG1 befinden sich in der Zelle an Stellen aktiven Wachstums. In Immun-Fluoreszenz Untersuchungen wurde eine partielle Ko-Lokalisation von POD6 und COT1 festgestellt. Diese Lokalisierung ist von Mikrotubuli Motorproteinen abhängig. LRG1 zeigt eine ähnliche, an Septen und der Hyphenspitze angereicherte, zelluläre Verteilung. Diese Lokalisation wurde sowohl mit gegen LRG1 gerichteten Antikörpern in Wildtyp-Zellen als auch für MYC9::LRG1 mit Antikörpern gegen das Myc-Epitop gefunden. Ein mit GFP fusioniertes LRG1-Protein findet sich in Abhängigkeit von intakten LIM Domänen und Aktin-Zytoskelett wachstumsabhängig als apikale Kappe, deren Größe mit der Wachstumsgeschwindigkeit korreliert. Ähnlich wie bei POD6 und COT1, wird auch diese Lokalisation wird von den Mikrotubuli Motoren Kinesin und Dynein beeinflusst. LRG1 beeinflusst die Aktivität verschiedener Effektorproteine von Rho1. Mutanten in lrg-1 sind unempfindlicher gegenüber dem Glucansynthase-Inhibitor Caspofungin und Doppelmutanten von lrg-1 mit einer hyperaktiven ß-1,3-Glucansynthase Mutante sind nicht lebensfähig. Die PKC-Inhibitoren Staurosporin und Cercosporamid unterdrücken die Wachstumssörungen von lrg-1 Mutanten, welche zudem empfindlicher gegenüber der Aktin depolymerisierenden Droge Latrunkulin A sind. Zudem wurde eine Unterdrückung der Wachstumsdefekte durch Expression des dominant negativ agierenden N-Terminus des Formins Bni gefunden. Die Untersuchung der Rho1 Wege in einer Mutante der NDR kinase cot-1 zeigte, dass diese Mutation keinen Einfluss auf die getesteten RHO1 Effektoren hat. Diese Experimente legen nahe, das LRG1 als spezifisches GAP für Rho1 verschiedene Effektorwege reguliert und das ein paralleler, von Rho1 unabhängiger, Signalweg zum polaren Wachstum erforderlich ist, in welchem COT1 und POD6 agieren

Case report: Eighteen nails gun shots in the head, thorax and abdomen but still conscious at admission: A challenging patient for the cardiac surgeon!

We report an unusual case of multiple penetrating cerebral, cardiac and abdominal injuries following a suicidal attempt using a nail gun. Successful treatment required several emergency procedures and resulted from a wise interdisciplinary management and timing of surgery

The calcium sensor Copine-6 regulates spine structural plasticity and learning and memory

Hippocampal long-term potentiation (LTP) represents the cellular response of excitatory synapses to specific patterns of high neuronal activity and is required for learning and memory. Here we identify a mechanism that requires the calcium-binding protein Copine-6 to translate the initial calcium signals into changes in spine structure. We show that Copine-6 is recruited from the cytosol of dendrites to postsynaptic spine membranes by calcium transients that precede LTP. Cpne6 knockout mice are deficient in hippocampal LTP, learning and memory. Hippocampal neurons from Cpne6 knockouts lack spine structural plasticity as do wild-type neurons that express a Copine-6 calcium mutant. The function of Copine-6 is based on its binding, activating and recruiting the Rho GTPase Rac1 to cell membranes. Consistent with this function, the LTP deficit of Cpne6 knockout mice is rescued by the actin stabilizer jasplakinolide. These data show that Copine-6 links activity-triggered calcium signals to spine structural plasticity necessary for learning and memory

Improving protein extraction and separation methods for investigating the metaproteome of anaerobic benzene communities within sediments

BTEX compounds such as benzene are frequent soil and groundwater contaminants that are easily biodegraded under oxic conditions by bacteria. In contrast, benzene is rather recalcitrant under anaerobic conditions. The analysis of anoxic degradation is often hampered by difficult sampling conditions, limited amounts of biomass and interference of matrix compounds with proteomic approaches. In order to improve the procedure for protein extraction we established a scheme consisting of the following steps: dissociation of cells from lava granules, cell lysis by ultrasonication and purification of proteins by phenol extraction. The 2D-gels revealed a resolution of about 240 proteins spots and the spot patterns showed strong matrix dependence, but still differences were detectable between the metaproteomes obtained after growth on benzene and benzoate. Using direct data base search as well as de novo sequencing approaches we were able to identify several proteins. An enoyl-CoA hydratase with cross species homology to Azoarcus evansii, is known to be involved in the anoxic degradation of xenobiotics. Thereby the identification confirmed that this procedure has the capacity to analyse the metaproteome of an anoxic living microbial community

Taxonomy and the Production of Semantic Phenotypes

Preprint of chapter appearing in "Studies on the Semantic Web: Volume 33: Application of Semantic Technology in Biodiversity Science"Taxonomists produce a myriad of phenotypic descriptions. Traditionally these are provided in terse (telegraphic) natural language. As seen in parallel within other fields of biology researchers are exploring ways to formalize parts of the taxonomic process so that aspects of it are more computational in nature. The currently used data formalizations, mechanisms for persisting data, applications, and computing approaches related to the production of semantic descriptions (phenotypes) are reviewed, they, and their adopters are limited in number. In order to move forward we step back and characterize taxonomists with respect to their typical workflow and tendencies. We then use these characteristics as a basis for exploring how we might create software that taxonomists will find intuitive within their cur- rent workflows, providing interface examples as thought experiments.NSF - DBI-1356381NSF 0956049https://deepblue.lib.umich.edu/bitstream/2027.42/148811/1/yoder_proof.pdfDescription of yoder_proof.pdf : Proof of book chapte

Calculation of partial isotope incorporation into peptides measured by mass spectrometry

<p>Abstract</p> <p>Background</p> <p>Stable isotope probing (SIP) technique was developed to link function, structure and activity of microbial cultures metabolizing carbon and nitrogen containing substrates to synthesize their biomass. Currently, available methods are restricted solely to the estimation of fully saturated heavy stable isotope incorporation and convenient methods with sufficient accuracy are still missing. However in order to track carbon fluxes in microbial communities new methods are required that allow the calculation of partial incorporation into biomolecules.</p> <p>Results</p> <p>In this study, we use the characteristics of the so-called 'half decimal place rule' (HDPR) in order to accurately calculate the partial¹³C incorporation in peptides from enzymatic digested proteins. Due to the clade-crossing universality of proteins within bacteria, any available high-resolution mass spectrometry generated dataset consisting of tryptically-digested peptides can be used as reference.</p> <p>We used a freely available peptide mass dataset from <it>Mycobacterium tuberculosis </it>consisting of 315,579 entries. From this the error of estimated versus known heavy stable isotope incorporation from an increasing number of randomly drawn peptide sub-samples (100 times each; no repetition) was calculated. To acquire an estimated incorporation error of less than 5 atom %, about 100 peptide masses were needed. Finally, for testing the general applicability of our method, peptide masses of tryptically digested proteins from <it>Pseudomonas putida </it>ML2 grown on labeled substrate of various known concentrations were used and¹³C isotopic incorporation was successfully predicted. An easy-to-use script <abbrgrp><abbr bid="B1">1</abbr></abbrgrp> was further developed to guide users through the calculation procedure for their own data series.</p> <p>Conclusion</p> <p>Our method is valuable for estimating¹³C incorporation into peptides/proteins accurately and with high sensitivity. Generally, our method holds promise for wider applications in qualitative and especially quantitative proteomics.</p

Geological interpretation of volcanism and segmentation of the Mariana back-arc spreading center between 12.7°N and 18.3°N

The relationships between tectonic processes, magmatism, and hydrothermal venting along ∼600 km of the slow-spreading Mariana back-arc between 12.7°N and 18.3°N reveal a number of similarities and differences compared to slow-spreading mid-ocean ridges. Analysis of the volcanic geomorphology and structure highlights the complexity of the back-arc spreading center. Here, ridge segmentation is controlled by large-scale basement structures that appear to predate back-arc rifting. These structures also control the orientation of the chains of cross-arc volcanoes that characterize this region. Segment-scale faulting is oriented perpendicular to the spreading direction, allowing precise spreading directions to be determined. Four morphologically distinct segment types are identified: dominantly magmatic segments (Type I); magmatic segments currently undergoing tectonic extension (Type II); dominantly tectonic segments (Type III); and tectonic segments currently undergoing magmatic extension (Type IV). Variations in axial morphology (including eruption styles, neovolcanic eruption volumes, and faulting) reflect magma supply, which is locally enhanced by cross-arc volcanism associated with N-S compression along the 16.5°N and 17.0°N segments. In contrast, cross-arc seismicity is associated with N-S extension and increased faulting along the 14.5°N segment, with structures that are interpreted to be oceanic core complexes—the first with high-resolution bathymetry described in an active back-arc basin. Hydrothermal venting associated with recent magmatism has been discovered along all segment types

Дослідження структури порушених відкритою розробкою земель й пошук шляхів вдосконалення рекультивації залишкових виробок кар'єрів

Стаття присвячена дослідженням структури порушених земель, на ділянках з видобутку корисних копалин відкритим способом. Наведено площі порушень земель при розробці основних видів корисних копалин. Проаналізовано ризики, що виникають із несвоєчасною рекультивацією земель гірничого відводу, а також від покинутих гірничих виробок старих кар'єрів. Паралельно розглянуті обсяги відходів гірничого виробництва та їх повторне використання в якості заповнювача для залишкових вироблених просторів кар'єрів.The article is devoted to the research of land violation indicators at the extraction of minerals by surface mining method. Data gives about the land violations area at the mining key minerals. Ana-lyzed the risks from the not-on-time reclamation of the mining clam and abandoned excavations of the old quarries. In parallel considered the volumes of mining wastes and their reuse as aggregate for filling residual spaces of surface mines.Статья посвящена исследованиям площадей нарушения земель, связанных с добычей полезных ископаемых открытым способом. Приведены площади нарушений земель при разработке основных видов полезных ископаемых. Проанализированы риски, представляемые несвоевременной рекультивацией земель горного отвода, а также заброшенными горными выработками старых карьеров. Параллельно рассмотрены объемы отходов горного производства и их повторное использование в качестве заполнителя для остаточных выработанных пространств карьеров