@phdthesis{Mueller2017,
  author    = {M{\"u}ller, Stephanie},
  title     = {Plant thermotolerance: The role of heat stress-induced triacylglycerols in \(Arabidopsis\) \(thaliana\)},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-152829},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2017},
  abstract  = {Plants are exposed to high temperature, especially during hot summer days. Temperatures are typically lowest in the morning and reach a maximum in the afternoon. Plants can tolerate and survive short-term heat stress even on hot summer days. A. thaliana seedlings have been reported to tolerate higher temperatures for different time periods, a phenomenon that has been termed basal thermotolerance. In addition, plants have the inherent capacity to acclimate to otherwise lethal temperatures. Arabidopsis thaliana seedlings acclimate at moderately elevated temperatures between 32-38° C. During heat acclimation, a genetically programmed heat shock response (HSR) is triggered that is characterized by a rapid activation of heat shock transcription factors (HSFs), which trigger a massive accumulation of heat shock proteins that are chiefly involved in protein folding and protection. Although the HSF-triggered heat-shock response is well characterized, little is known about the metabolic adjustments during heat stress. The aim of this work was to get more insight into heat-responsive metabolism and its importance for thermotolerance. In order to identify the response of metabolites to elevated temperatures, global metabolite profiles of heat-acclimated and control seedlings were compared. Untargeted metabolite analyses revealed that levels of polyunsaturated triacylglycerols (TG) rapidly increase during heat acclimation. TG accumulation was found to be temperature-dependent in a temperature range from 32-50° C (optimum at 42° C). Heat-induced TG accumulation was localized in extra-chloroplastic compartments by chloroplast isolation as well as by fluorescence microscopy of A. thaliana cell cultures. Analysis of mutants deficient in all four HSFA1 master regulator genes or the HSFA2 gene revealed that TG accumulation occurred independently to HSF. Moreover, the TG response was not limited to heat stress since drought and salt stress (but not short-term osmotic, cold and high light stress) also triggered an accumulation of TGs. In order to reveal the origin of TG synthesis, lipid analysis was carried out. Heat-induced accumulation of TGs does not derive from massive de novo fatty acid (FA) synthesis. On the other hand, lipidomic analyses of A. thaliana seedlings indicated that polyunsaturated FA from thylakoid galactolipids are incorporated into cytosolic TGs during heat stress. This was verified by lipidomic analyses of A. thaliana fad7/8 transgenic seedlings, which displayed altered FA compositions of plastidic lipids. In addition, wild type A. thaliana seedlings displayed a rapid conversion of plastidic monogalactosyldiacylglycerols (MGDGs) into oligogalactolipids, acylated MGDGs and diacylglycerols (DGs). For TG synthesis, DG requires a FA from the acyl CoA pool or phosphatidylcholine (PC). Seedlings deficient in phospholipid:diacylglycerol acyltransferase1 (PDAT1) were unable to accumulate TGs following heat stress; thus PC appears to be the major FA donor for TGs during heat treatment. These results suggest that TG and oligogalactolipid accumulation during heat stress is driven by post-translationally regulated plastid lipid metabolism. TG accumulation following heat stress was found to increase basal thermotolerance. Pdat1 mutant seedlings were more sensitive to severe heat stress without prior acclimatization, as revealed by a more dramatic decline of the maximum efficiency of PSII and lower survival rate compared to wild type seedlings. In contrast, tgd1 mutants over-accumulating TGs and oligogalactolipids displayed a higher basal thermotolerance compared to wild type seedlings. These results therefore suggest that accumulation of TGs increases thermotolerance in addition to the genetically encoded heat shock response.},
  subject      = {Triglyceride},
  language  = {en}
}
@phdthesis{Horn2017,
  author    = {Horn, Hannes},
  title     = {Analysis and interpretation of (meta-)genomic data from host-associated microorganisms},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-152035},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2017},
  abstract  = {Host-microbe interactions are the key to understand why and how microbes inhabit specific environments. With the scientific fields of microbial genomics and metagenomics, evolving on an unprecedented scale, one is able to gain insights in these interactions on a molecular and ecological level. The goal of this PhD thesis was to make (meta-)genomic data accessible, integrate it in a comparative manner and to gain comprehensive taxonomic and functional insights into bacterial strains and communities derived from two different environments: the phyllosphere of Arabidopsis thaliana and the mesohyl interior of marine sponges. This thesis focused first on the de novo assembly of bacterial genomes. A 5-step protocol was developed, each step including a quality control. The examination of different assembly software in a comparative way identified SPAdes as most suitable. The protocol enables the user to chose the best tailored assembly. Contamination issues were solved by an initial filtering of the data and methods normally used for the binning of metagenomic datasets. This step is missed in many published assembly pipelines. The described protocol offers assemblies of high quality ready for downstream analysis. Subsequently, assemblies generated with the developed protocol were annotated and explored in terms of their function. In a first study, the genome of a phyllosphere bacterium, Williamsia sp. ARP1, was analyzed, offering many adaptions to the leaf habitat: it can deal with temperature shifts, react to oxygen species, produces mycosporins as protection against UV-light, and is able to uptake photosynthates. Further, its taxonomic position within the Actinomycetales was infered from 16S rRNA and comparative genomics showing the close relation between the genera Williamsia and Gordonia. In a second study, six sponge-derived actinomycete genomes were investigated for secondary metabolism. By use of state-of-the-art software, these strains exhibited numerous gene clusters, mostly linked to polykethide synthases, non-ribosomal peptide synthesis, terpenes, fatty acids and saccharides. Subsequent predictions on these clusters offered a great variety of possible produced compounds with antibiotic, antifungal or anti-cancer activity. These analysis highlight the potential for the synthesis of natural products and the use of genomic data as screening toolkit. In a last study, three sponge-derived and one seawater metagenomes were functionally compared. Different signatures regarding the microbial composition and GC-distribution were observed between the two environments. With a focus on bacerial defense systems, the data indicates a pronounced repertoire of sponge associated bacteria for bacterial defense systems, in particular, Clustered Regularly Interspaced Short Palindromic Repeats, restriction modification system, DNA phosphorothioation and phage growth limitation. In addition, characterizing genes for secondary metabolite cluster differed between sponge and seawater microbiomes. Moreover, a variety of Type I polyketide synthases were only found within the sponge microbiomes. With that, metagenomics are shown to be a useful tool for the screening of secondary metabolite genes. Furthermore, enriched defense systems are highlighted as feature of sponge-associated microbes and marks them as a selective trait.},
  subject      = {Bakterien},
  language  = {en}
}
@phdthesis{Kums2017,
  author    = {Kums, Juliane},
  title     = {Entwicklung und Charakterisierung von \(Gaussia\) \(princeps\) Luziferase-Antik{\"o}rper-Fusionsproteinen},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-146777},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2017},
  abstract  = {Antik{\"o}rper, die Oberfl{\"a}chenantigene erkennen, sind sowohl in der Diagnostik als auch in der Therapie verschiedener Erkrankungen von enormer Bedeutung. Damit Antik{\"o}rper in diesen Bereichen eingesetzt werden k{\"o}nnen, ist es sehr wichtig, dass die Interaktion eines Antik{\"o}rpers oder auch eines Antik{\"o}rperkonjugats mit seinem Antigen oder Fc-Rezeptoren ausreichend charakterisiert wird. Hierf{\"u}r werden meist zellfreie Verfahren angewandt, wie die isotherme Titrationskalorimetrie oder die Oberfl{\"a}chenplasmonenresonanzspektroskopie. Diese unterliegen verschiedenen Limitationen, beispielsweise der Verf{\"u}gbarkeit von rekombinantem Antigen. Vor allem aber werden zellul{\"a}re Einfl{\"u}sse, die die Bindungseigenschaften der Antik{\"o}rper beeinflussen, nicht ber{\"u}cksichtigt. Aber auch die derzeit angewandten Verfahren f{\"u}r zellul{\"a}re Bindungsstudien k{\"o}nnen problematisch sein, da sie meist auf Antik{\"o}rpern basieren, die biochemisch markiert worden sind, was zu funktionellen Beeintr{\"a}chtigungen f{\"u}hren kann. Außerdem zeigen solche Antik{\"o}rper h{\"a}ufig keine einheitliche St{\"o}chiometrie der jeweiligen Reporterstoffe und die Reproduzierbarkeit des Markierungsverfahrens ist in den meisten F{\"a}llen nicht gew{\"a}hrleistet. Positionsspezifische Markierungen sind jedoch vergleichsweise sehr aufwendig. Um die genannten Probleme zu umgehen, wurden in der vorliegenden Arbeit am Beispiel des Fn14-spezifischen Antik{\"o}rpers 18D1 Antik{\"o}rper-Fusionsproteine hergestellt und charakterisiert, die an verschiedenen Positionen genetisch mit der Gaussia princeps Luziferase (GpL) fusioniert worden sind. Dabei zeigte sich, dass die Positionierung der Luziferase am C-Terminus der leichten Kette des Antik{\"o}rpers (GpL(CT-LC)) die Bindungseigenschaften der GpL-18D1-IgG1-Fusionsproteine an Fn14 und an die verschiedenen Fcγ-Rezeptoren (FcγR) nicht oder nur in geringem Umfang beeinflusst. Auch die agonistische Aktivit{\"a}t der GpL-18D1-IgG1-Fusionsproteine, welche abh{\"a}ngig ist von der Oligomerisierung {\"u}ber Protein G oder der FcγR-Bindung, wurde durch die GpL-Markierung nicht wesentlich beeinflusst. Diese Ergebnisse ließen sich am Bespiel von 18D1 ebenfalls auf die dimeren Antik{\"o}rper-Isotypen IgG2, mIgG1 und mIgG2A {\"u}bertragen. GpL-Fusionsproteine der Antik{\"o}rper E09-IgG1 (CD95-spezifisch), G28.5-IgG1 (CD40-spezifisch) und BHA10-IgG1 (LTβR-spezifisch) zeigten gleichfalls keine gravierenden Ver{\"a}nderungen der Bindungseigenschaften oder den funktionellen Eigenschaften, was f{\"u}r eine breite Anwendbarkeit von GpL-Antik{\"o}rper-Fusionsproteinen spricht. Zusammenfassend betrachtet zeigen die hier pr{\"a}sentierten Ergebnisse, dass die genetische Fusion der Gaussia princeps Luziferase an das C-terminale Ende der leichten Antik{\"o}rperkette eine sehr gute M{\"o}glichkeit darstellt, Antigen-Antik{\"o}rper-Interaktionen zu charakterisieren ohne dabei mit den Eigenschaften des Antik{\"o}rpers zu interferieren. Dabei besticht dieser Ansatz im Vergleich zu anderen g{\"a}ngigen Verfahren durch seine Reproduzierbarkeit, eine einfache Handhabung, geringe Kosten und eine extrem hohe Sensitivit{\"a}t. Außerdem k{\"o}nnte dieses Antik{\"o}rper-Fusionsproteinformat zuk{\"u}nftig auch in vielen Bereichen als Tracer eingesetzt werden mit dem Vorteil, dass keinerlei Radioaktivit{\"a}t ben{\"o}tigt werden w{\"u}rde.},
  subject      = {Luciferasen},
  language  = {de}
}
@phdthesis{Gutermuth2017,
  author    = {Gutermuth, Timo},
  title     = {Identifizierung und Charakterisierung des Signalweges zur Aktivierung von Anionenkan{\"a}len w{\"a}hrend des Pollenschlauchwachstums},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-139232},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2017},
  abstract  = {Pollenschl{\"a}uche sind ein Modellsystem zur Untersuchung pflanzlicher Wachstumsprozesse. Zellwachstum in Pollenschl{\"a}uchen zeichnet sich durch den gerichteten Transport und Fusion von Vesikeln mit der apikalen Zellmembran des Pollenschlauchs aus. Der Vesikeltransport erfolgt entlang des Pollenschlauchs durch Aktin-Filamente bis an die Organell- und Zytoskelett-freie apikale Zone, wo sich die Vesikel sammeln und in oszillierenden Wachstumssch{\"u}ben mit der apikalen Zellmembran fusionieren (Yang et al., 1998; Zonia et al., 2001, Gu et al., 2005; Chen et al., 2003; Gu et al., 2005; de Graaf et al., 2005; Lee et al., 2008; Cheung et al., 2010; Quin und Yang et al., 2011). Die polaren Wachstumsprozesse des Pollenschlauches sind an ein Ionenflussmuster gekoppelt, welches durch den Einsatz der Vibrating Probe-Technik zeitlich aufgel{\"o}st werden konnten. Es konnte ein zeitversetzter oszillierender Einstrom von Calcium, Kalium und Protonen sowie der zeitgleich mit den Wachstumssch{\"u}ben auftretende oszillierende Ausstrom von Chlorid aus der Pollenschlauchspitze nachgewiesen werden (K{\"u}htreiber und Jaffe et al., 1990; Holdaway-Clarke et al., 1997; Feijo et al., 1999, Messerli et al., 1999, Zonia et al., 2001). Die Inhibierung des Chloridausstroms resultiert in einem sofortigen Wachstumsstopp und verdeutlicht die Notwendigkeit des Anionenausstroms f{\"u}r das polare Zellwachstum in Pollenschl{\"a}uchen (Breygina et al., 2009). Durch die in dieser Arbeit durchgef{\"u}hrten Experimente konnten die an dem Anionenausstrom beteiligten Anionenkan{\"a}le, sowie deren Ca2+-abh{\"a}ngigen regulatorischen Komponenten identifiziert und mit Hilfe der TEVC-Technik elektrophysiologisch an intakten Arabidopsis thaliana-Pollenschl{\"a}uchen charakterisiert werden. Weiterhin konnte die physiologische Rolle der f{\"u}r den Anionenausstrom verantwortlichen Kan{\"a}le auf das polare Zellwachstum in Arabidopsis thaliana Pollenschl{\"a}uchen nachgewiesen werden. Durch Transkriptionsanalysen wurde die Expression des S-Typ-Anionenkanals SLAH3 sowie der R-Typ-Anionenkan{\"a}le ALMT12, ALMT13 und ALMT14 in Arabidopsis thaliana Pollenschl{\"a}uchen belegt und deren transkriptionelle Regulation durch die Anionenkonzentration und Komposition des Keimungsmediums nachgewiesen werden. Eine elektrophysiologische Charakterisierung an intakten Arabidopsis thaliana Pollenschl{\"a}uchen konnte sowohl einen Anstieg der SLAH3 vermittelten S-Typ-Str{\"o}me, als auch ALMT12-, ALMT13- und ALMT14 vermittelte R Typ-Anionenstr{\"o}me bei steigenden Anionenkonzentrationen im Keimungsmedium nachweisen. Die Charakterisierung der Verlustmutanten von SLAH3, ALMT12, ALMT13 und ALMT14 resultierte in einer Abnahme des Anionenausstroms und einer Reduktion des L{\"a}ngenwachstums der getesteten Mutanten. Es konnten ebenfalls die regulatorischen Komponenten der Signalkette zur Anionenkanalaktivierung identifiziert werden. Die Aktivierung von SLAH3 und ALMT12 durch die Calcium-abh{\"a}ngigen Kinasen CPK2, CPK20 und CPK6 aus Arabidopsis thaliana Pollenschl{\"a}uchen konnte mittels einer Kombination von elektrophysiologischen- und molekularbiologischen Techniken nachgewiesen werden. Somit wurden nicht nur die f{\"u}r den Anionenausstrom verantwortlichen Anionenkan{\"a}le identifiziert, sondern auch die Signalkette zu deren Aktivierung durch spitzenlokalisierte Calcium-abh{\"a}ngige Kinasen aufgekl{\"a}rt werden. Diese Signalkaskade f{\"u}hrt ebenfalls durch die artifizielle Erh{\"o}hung der zytoplasmatischen Calciumkonzentration durch das Calcium-Ionophor A23187 zu einem Anstieg des S Typ- und R Typ Anionenkanalaktivit{\"a}t in Arabidopsis thaliana-Pollenschl{\"a}uchen. Eine intensivere Charakterisierung des entdeckten Calcium-vermittelten Anionenausstroms erfolgte am transgenen pLat52-Chlorid-Sensor bzw. an YC3.6 Tabak Pollenschl{\"a}uchen durch die Kombination von TEVC-Technik und Fluoreszensmikroskopie. Dies erm{\"o}glichte die simultane Messung der zytoplasmatischen Calcium- bzw. Chloridkonzentration in Nicotiana tabacum Pollenschl{\"a}uchen bei gleichzeitiger Ableitung der Ganzzellstr{\"o}me. Die elektrophysiologische und fluoreszenzmikroskopische Charakterisierung erbrachte erstmals den Nachweis f{\"u}r eine exklusive Lokalisation von hyperpolarisations-aktivierten Calciumkan{\"a}len in der Pollenschlauchspitze, welche sich durch die Verwendung der TEVC-Technik gezielt aktivieren ließen. Diese Aktivierung der spitzenlokalisierten Calciumkan{\"a}le induziert den Anionenausstrom durch den Anstieg der apikalen Calciumkonzentration. Die Inhibierung der Calciumkan{\"a}le durch den Calciumkanalblocker Lanthan f{\"u}hrt zu einem vollst{\"a}ndigen Verlust des Calciumeinstroms und des daraus resultierenden Anioneneinstroms. Durch die Inhibierung der Calciumkan{\"a}le kommt es gleichzeitig zu einer Akkumulation von Chlorid in der apikalen Zone, die zum Anschwellen der Pollenschlauchspitze f{\"u}hrt. Die Inhibierung der Anionenkan{\"a}le durch Niflums{\"a}ure hat hingegen keinen Einfluss auf den spitzenlokalisierten Calciumeinstrom, sondern reduziert nur den gemessenen Anionenausstrom. Somit wird ein kausaler Zusammenhang zwischen der Erh{\"o}hung der apikalen Ca2+-Konzentration und einer Anionenkanalaktivierung weiter verdeutlicht. Durch die Anwendung der TEVC-Technik an intakten Pollenschl{\"a}uchen konnten erstmals Aktionspotenzial {\"a}hnliche Depolarisierungstransienten, welche sich auf die apikale Zone des Pollenschlauchs beschr{\"a}nken und zeitgleich mit dem Anionenausstrom stattfinden, nachgewiesen werden. Durch diese Arbeit kann erstmals ein Modell des Calcium-vermittelten oszillierenden Anionenausstroms aus der Pollenschlauchspitze aufgestellt werden. Dieses verkn{\"u}pft die Regulation der beteiligten R-Typ-Anionenkan{\"a}le ALMT12, ALMT13 und ALMT14 und des S-Typ-Anionenkanals SLAH3 durch die Calcium-abh{\"a}ngigen Kinasen CPK2, CPK20 und CPK6 mit dem spitzenlokalisierten oszillierenden Calciumeinstrom. Das Modell verdeutlicht die physiologische Bedeutung des simultanen Ca2+-Ein- und Anionenausstroms f{\"u}r das polare Zellwachstum von Pollenschl{\"a}uchen.  },
  subject      = {Pollenschlauch},
  language  = {de}
}
@phdthesis{Gehring2017,
  author    = {Gehring, Jennifer},
  title     = {Functional analysis of the latrophilin homolog dCirl in Drosophila melanogaster},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-101061},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2017},
  abstract  = {Latrophilin, alternatively named calcium-independent receptor of α-latrotoxin (CIRL), resembles a prototype of the adhesion class G-protein coupled receptors (GPCRs). Initially identified as a high-affinity receptor for α-latrotoxin, a component of the black widow spider, latrophilins are now associated with various distinct functions, such as synaptic exocytosis, tissue polarity and fertility (Tobaben et al., 2002; Langenhan et al., 2009; Promel et al., 2012). Despite these exploratory efforts the precise subcellular localisation as well as the endogenous ligand of CIRL still remains elusive. In this work genetic experiments, imaging approaches and behavioural studies have been used to unravel the localisation and physiological function of the latrophilin homolog dCirl in Drosophila melanogaster. Containing only one latrophilin homolog together with its genetic accessibility and well-established transgenic approaches, Drosophila seemed an ideally suited model organism. The present study showed that dCirl is widely expressed in the larval central nervous system including moto- and sensory neurons. Further, this work revealed that removal of the latrophilin homolog does not greatly affect synaptic transmission but it seems that aspects of the postsynaptic structural layout are controlled by dCIRL in the fruit fly. Additionally, dCirl expression at the transcriptional level was confirmed in larval and adult chordotonal organs, specialised mechanosensors implicated in proprioception (Eberl, 1999). Expression of dCIRL at the protein level could not yet been confirmed in moto- and sensory neurons likely due to low endogenous expression. However, behavioural studies using dCirl knockout mutant larvae indicated a putative mechanosensory function of dCIRL regarding touch sensitivity and locomotion behaviour. The second part of this thesis presents a strategy to examine interactions between several presynaptic proteins in living cells. The attempt described in this work is based on the discovery that GFP when split into two non-fluorescent fragments can form a fluorescent complex. The association of the fragments can be facilitated by fusing them to two proteins that interact with each other. Therefore, the split GFP method enables direct visualization of synaptic protein interactions in living cells. In initial experiments I could show that full length reporter protein fusions with n-Synaptobrevin (n-Syb), Synaptotagmin (Syt) and Syntaxin (Syx) allow expression in Drosophila and confirmed that fusion to either end of each synaptic protein did not impair expression or influence the viability of transgenic flies. Further, transgenes containing protein fusions of Syx, Syt, and n-Syb with split GFP fragments were established in previous studies (Gehring, 2010). The present work characterises the interaction of these protein fusions during different stages of synaptic vesicle turnover at active zones such as synaptic vesicle docking at the presynaptic membrane and vesicle fusion. These results suggest that the spGFP assay seems only partly suitable for resolving fast and transient protein-protein interactions at larval Drosophila active zones in vivo.},
  subject      = {Taufliege},
  language  = {en}
}
@phdthesis{Gao2017,
  author    = {Gao, Shiqiang},
  title     = {Characterizing new photoreceptors to expand the Optogenetic toolbox},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-112941},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2017},
  abstract  = {Optogenetics is a method to control the cell activity with light by expression of a natural or engineered photoreceptor via genetic modification technology. Optogenetics early success came with the light-gated cation channel "Channelrhodopsin-2" in neurons and expanded from neuroscience to other research fields such as cardiac research and cell signaling, also due to the enrichment by new photoreceptors. In this study, I focus on searching and characterizing new photoreceptors to expand the optogenetic tool box. In this work I characterize three newly discovered microbial rhodopsins and some engineered mutants of them. The first rhodopsin is a proton pump from the diatom Fragilariopsis cylindrus, Fragilariopsis Rhodopsin or abbreviated: FR. I cloned the full-length FR and proved it to be a light-activated proton pump with high efficacy in comparison to Bacteriorhodopsin (BR). During this study, I also developed a new method to improve the plasma membrane targeting of several microbial rhodopsins. I also obtained a FR mutant (channel-like FR or chFR) which behaves like a light-gated proton channel. FR can be used for optogenetic hyperpolarization or alkalization of a cell while the chFR could be used for depolarization or lowering of the cellular pH. The induction of FR expression under iron-limited conditions in the diatom indicated an alternative energy generation mechanism of F. cylindrus when iron-containing enzymes are scarce. I then characterized a new microbial rhodopsin with novel light-regulated Guanylyl Cyclase (GC) activity. This rhodopsin guanylyl cyclase from the fungus Blastocladiella emersonii (B.e. CyclaseOpsin or BeCyclOp) has been proven by me to be an efficient light-gated GC with high specificity and fast kinetics. BeCyclOp also has a novel structure with eight transmembrane helices, containing a long cytosolic N-terminus which participates in the tight regulation of the GC activity. In collaboration with Prof. Alexander Gottschalk (Univ. Frankfurt/M.), BeCyclOp has been tested in muscle cells and sensory neurons of Caenorhabditis elegans and proven to be a powerful optogenetic tool in a living animal. I also generated a BeCyclOp mutant with enhanced light sensitivity. Already more than ten years ago, guanylyl cyclase rhodopsins were suggested to exist in Chlamydomonas reinhardtii by analyzing genomic sequence data. But until now no functional proof existed. By further cloning and sequencing I discovered such a rhodopsin with light-regulated guanylyl cyclase activity. This functional Cyclaseopsin (COP6c) is quite different to BeCyclOp, as it was proven to be a light-inhibited GC. Cop6c is much larger than BeCyclOp with a His-Kinase and a response regulator domain between the rhodopsin and the cyclase domain. I also introduced a new strategy for generating optogenetic tools by fusing the photoactivated adenylyl cyclase bPAC to two different CNG channels. These new tools function via light-gated cAMP production and subsequent CNG channel activation. These tools combined the properties of bPAC (highly sensitive to blue light) and CNG channels (high single-channel conductance and high Ca2+ permeability), as demonstrated by expression in Xenopus oocytes. As a further benefit the fusing of bPAC to CNG channels leads to a bPAC with a more than tenfold reduced dark activity which is a valuable improvement for bPAC itself as an optogenetic tool.},
  subject      = {Photorezeptor},
  language  = {en}
}
@article{LaglerElMeseryKuebleretal.2017,
  author    = {Lagler, Charlotte and El-Mesery, Mohamed and K{\"u}bler, Alexander Christian and M{\"u}ller-Richter, Urs Dietmar Achim and St{\"u}hmer, Thorsten and Nickel, Joachim and M{\"u}ller, Thomas Dieter and Wajant, Harald and Seher, Axel},
  title     = {The anti-myeloma activity of bone morphogenetic protein 2 predominantly relies on the induction of growth arrest and is apoptosis-independent},
  series = {PLoS ONE},
  volume    = {12},
  journal   = {PLoS ONE},
  number    = {10},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-158993},
  pages     = {e0185720},
  year      = {2017},
  abstract  = {Multiple myeloma (MM), a malignancy of the bone marrow, is characterized by a pathological increase in antibody-producing plasma cells and an increase in immunoglobulins (plasmacytosis). In recent years, bone morphogenetic proteins (BMPs) have been reported to be activators of apoptotic cell death in neoplastic B cells in MM. Here, we use bone morphogenetic protein 2 (BMP2) to show that the "apoptotic" effect of BMPs on human neoplastic B cells is dominated by anti-proliferative activities and cell cycle arrest and is apoptosis-independent. The anti-proliferative effect of BMP2 was analysed in the human cell lines KMS12-BM and L363 using WST-1 and a Coulter counter and was confirmed using CytoTox assays with established inhibitors of programmed cell death (zVAD-fmk and necrostatin-1). Furthermore, apoptotic activity was compared in both cell lines employing western blot analysis for caspase 3 and 8 in cells treated with BMP2 and FasL. Additionally, expression profiles of marker genes of different cell death pathways were analysed in both cell lines after stimulation with BMP2 for 48h using an RT-PCR-based array. In our experiments we observed that there was rather no reduction in absolute cell number, but cells stopped proliferating following treatment with BMP2 instead. The time frame (48-72 h) after BMP2 treatment at which a reduction in cell number is detectable is too long to indicate a directly BMP2-triggered apoptosis. Moreover, in comparison to robust apoptosis induced by the approved apoptotic factor FasL, BMP2 only marginally induced cell death. Consistently, neither the known inhibitor of apoptotic cell death zVAD-fmk nor the necroptosis inhibitor necrostatin-1 was able to rescue myeloma cell growth in the presence of BMP2.},
  language  = {en}
}
@article{ChengOthmanStopperetal.2017,
  author    = {Cheng, Cheng and Othman, Eman M. and Stopper, Helga and Edrada-Ebel, RuAngelie and Hentschel, Ute and Abdelmohsen, Usama Ramadan},
  title     = {Isolation of petrocidin A, a new cytotoxic cyclic dipeptide from the marine sponge-derived bacterium \(Streptomyces\) sp. SBT348},
  series = {Marine Drugs},
  volume    = {15},
  journal   = {Marine Drugs},
  number    = {12},
  doi       = {10.3390/md15120383},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-172644},
  year      = {2017},
  abstract  = {A new cyclic dipeptide, petrocidin A (\(\textbf{1}\)), along with three known compounds—2,3-dihydroxybenzoic acid (\(\textbf{2}\)), 2,3-dihydroxybenzamide (\(\textbf{3}\)), and maltol (\(\textbf{4}\))—were isolated from the solid culture of \(Streptomyces\) sp. SBT348. The strain \(Streptomyces\) sp. SBT348 had been prioritized in a strain collection of 64 sponge-associated actinomycetes based on its distinct metabolomic profile using liquid chromatography/high-resolution mass spectrometry (LC-HRMS) and nuclear magnetic resonance (NMR). The absolute configuration of all α-amino acids was determined by HPLC analysis after derivatization with Marfey's reagent and comparison with commercially available reference amino acids. Structure elucidation was pursued in the presented study by mass spectrometry and NMR spectral data. Petrocidin A (\(\textbf{1}\)) and 2,3-dihydroxybenzamide (\(\textbf{3}\)) exhibited significant cytotoxicity towards the human promyelocytic HL-60 and the human colon adenocarcinoma HT-29 cell lines. These results demonstrated the potential of sponge-associated actinomycetes for the discovery of novel and pharmacologically active natural products.},
  language  = {en}
}
@article{SalkerSinghZengetal.2017,
  author    = {Salker, Madhuri S. and Singh, Yogesh and Zeng, Ni and Chen, Hong and Zhang, Shaqiu and Umbach, Anja T. and Fakhri, Hajar and Kohlhofer, Ursula and Quintanilla-Martinez, Leticia and Durairaj, Ruban R. Peter and Barros, Flavio S. V. and Vrljicak, Pavle and Ott, Sascha and Brucker, Sara Y. and Wallwiener, Diethelm and Madunić, Ivana Vrhovac and Breljak, Davorka and Sabolić, Ivan and Koepsell, Hermann and Brosens, Jan J. and Lang, Florian},
  title     = {Loss of endometrial sodium glucose cotransporter SGLT1 is detrimental to embryo survival and fetal growth in pregnancy},
  series = {Scientific Reports},
  volume    = {7},
  journal   = {Scientific Reports},
  doi       = {10.1038/s41598-017-11674-3},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-173814},
  year      = {2017},
  abstract  = {Embryo implantation requires a hospitable uterine environment. A key metabolic change that occurs during the peri-implantation period, and throughout early pregnancy, is the rise in endometrial glycogen content. Glycogen accumulation requires prior cellular uptake of glucose. Here we show that both human and murine endometrial epithelial cells express the high affinity Na\(^+\)-coupled glucose carrier SGLT1. Ussing chamber experiments revealed electrogenic glucose transport across the endometrium in wild type (\(Slc5a1^{+/+}\)) but not in SGLT1 defcient (\(Slc5a1^{-/-}\)) mice. Endometrial glycogen content, litter size and weight of offspring at birth were signifcantly lower in \(Slc5a1^{-/-}\) mice. In humans, \(SLC5A1\) expression was upregulated upon decidualization of primary endometrial stromal cells. Endometrial \(SLC5A1\) expression during the implantation window was attenuated in patients with recurrent pregnancy loss when compared with control subjects. Our fndings reveal a novel mechanism establishing adequate endometrial glycogen stores for pregnancy. Disruption of this histiotrophic pathway leads to adverse pregnancy outcome.},
  language  = {en}
}
@article{BalasubramanianOthmanKampiketal.2017,
  author    = {Balasubramanian, Srikkanth and Othman, Eman M. and Kampik, Daniel and Stopper, Helga and Hentschel, Ute and Ziebuhr, Wilma and Oelschlaeger, Tobias A. and Abdelmohsen, Usama R.},
  title     = {Marine sponge-derived Streptomyces sp SBT343 extract inhibits staphylococcal biofilm formation},
  series = {Frontiers in Microbiology},
  volume    = {8},
  journal   = {Frontiers in Microbiology},
  doi       = {10.3389/fmicb.2017.00236},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-171844},
  year      = {2017},
  abstract  = {Staphylococcus epidermidis and Staphylococcus aureus are opportunistic pathogens that cause nosocomial and chronic biofilm-associated infections. Indwelling medical devices and contact lenses are ideal ecological niches for formation of staphylococcal biofilms. Bacteria within biofilms are known to display reduced susceptibilities to antimicrobials and are protected from the host immune system. High rates of acquired antibiotic resistances in staphylococci and other biofilm-forming bacteria further hamper treatment options and highlight the need for new anti-biofilm strategies. Here, we aimed to evaluate the potential of marine sponge-derived actinomycetes in inhibiting biofilm formation of several strains of S. epidermidis, S. aureus, and Pseudomonas aeruginosa. Results from in vitro biofilm-formation assays, as well as scanning electron and confocal microscopy, revealed that an organic extract derived from the marine sponge-associated bacterium Streptomyces sp. SBT343 significantly inhibited staphylococcal biofilm formation on polystyrene, glass and contact lens surfaces, without affecting bacterial growth. The extract also displayed similar antagonistic effects towards the biofilm formation of other S. epidermidis and S. aureus strains tested but had no inhibitory effects towards Pseudomonas biofilms. Interestingly the extract, at lower effective concentrations, did not exhibit cytotoxic effects on mouse fibroblast, macrophage and human corneal epithelial cell lines. Chemical analysis by High Resolution Fourier Transform Mass Spectrometry (HRMS) of the Streptomyces sp. SBT343 extract proportion revealed its chemical richness and complexity. Preliminary physico-chemical characterization of the extract highlighted the heat-stable and non-proteinaceous nature of the active component(s). The combined data suggest that the Streptomyces sp. SBT343 extract selectively inhibits staphylococcal biofilm formation without interfering with bacterial cell viability. Due to absence of cell toxicity, the extract might represent a good starting material to develop a future remedy to block staphylococcal biofilm formation on contact lenses and thereby to prevent intractable contact lens-mediated ocular infections.},
  language  = {en}
}