TY  - JOUR
A1  - Mercier, Rebecca
A1  - Wolmarans, Annemarie
A1  - Schubert, Jonathan
A1  - Neuweiler, Hannes
A1  - Johnson, Jill L.
A1  - LaPointe, Paul
T1  - The conserved NxNNWHW motif in Aha-type co-chaperones modulates the kinetics of Hsp90 ATPase stimulation
JF  - Nature Communications
N2  - Hsp90 is a dimeric molecular chaperone that is essential for the folding and activation of hundreds of client proteins. Co-chaperone proteins regulate the ATP-driven Hsp90 client activation cycle. Aha-type co-chaperones are the most potent stimulators of the Hsp90 ATPase activity but the relationship between ATPase regulation and in vivo activity is poorly understood. We report here that the most strongly conserved region of Aha-type co-chaperones, the N terminal NxNNWHW motif, modulates the apparent affinity of Hsp90 for nucleotide substrates. The ability of yeast Aha-type co-chaperones to act in vivo is ablated when the N terminal NxNNWHW motif is removed. This work suggests that nucleotide exchange during the Hsp90 functional cycle may be more important than rate of catalysis.
KW  - biophysics
KW  - cell growth
KW  - chaperones
KW  - enzymes
Y1  - 2019
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-224007
VL  - 10
ER  - 
TY  - JOUR
A1  - Lübcke, Paul M.
A1  - Ebbers, Meinolf N. B.
A1  - Volzke, Johann
A1  - Bull, Jana
A1  - Kneitz, Susanne
A1  - Engelmann, Robby
A1  - Lang, Hermann
A1  - Kreikemeyer, Bernd
A1  - Müller-Hilke, Brigitte
T1  - Periodontal treatment prevents arthritis in mice and methotrexate ameliorates periodontal bone loss
JF  - Scientific Reports
N2  - Recent studies indicate a causal relationship between the periodontal pathogen P. gingivalis and rheumatoid arthritis involving the production of autoantibodies against citrullinated peptides. We therefore postulated that therapeutic eradication P. gingivalis may ameliorate rheumatoid arthritis development and here turned to a mouse model in order to challenge our hypothesis. F1 (DBA/1 x B10.Q) mice were orally inoculated with P. gingivalis before collagen-induced arthritis was provoked. Chlorhexidine or metronidazole were orally administered either before or during the induction phase of arthritis and their effects on arthritis progression and alveolar bone loss were compared to intraperitoneally injected methotrexate. Arthritis incidence and severity were macroscopically scored and alveolar bone loss was evaluated via microcomputed tomography. Serum antibody titres against P. gingivalis were quantified by ELISA and microbial dysbiosis following oral inoculation was monitored in stool samples via microbiome analyses. Both, oral chlorhexidine and metronidazole reduced the incidence and ameliorated the severity of collagen-induced arthritis comparable to methotrexate. Likewise, all three therapies attenuated alveolar bone loss. Relative abundance of Porphyromonadaceae was increased after oral inoculation with P. gingivalis and decreased after treatment. This is the first study to describe beneficial effects of non-surgical periodontal treatment on collagen-induced arthritis in mice and suggests that mouthwash with chlorhexidine or metronidazole may also be beneficial for patients with rheumatoid arthritis and a coexisting periodontitis. Methotrexate ameliorated periodontitis in mice, further raising the possibility that methotrexate may also positively impact on the tooth supporting tissues of patients with rheumatoid arthritis.
KW  - rheumatic diseases
KW  - rheumatology
Y1  - 2019
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-237355
VL  - 9
ER  - 
TY  - JOUR
A1  - Lu, Yuan
A1  - Boswell, Wiliam
A1  - Boswell, Mikki
A1  - Klotz, Barbara
A1  - Kneitz, Susanne
A1  - Regneri, Janine
A1  - Savage, Markita
A1  - Mendoza, Cristina
A1  - Postlethwait, John
A1  - Warren, Wesley C.
A1  - Schartl, Manfred
A1  - Walter, Ronald B.
T1  - Application of the Transcriptional Disease Signature (TDSs) to Screen Melanoma-Effective Compounds in a Small Fish Model
JF  - Scientific Reports
N2  - Cell culture and protein target-based compound screening strategies, though broadly utilized in selecting candidate compounds, often fail to eliminate candidate compounds with non-target effects and/or safety concerns until late in the drug developmental process. Phenotype screening using intact research animals is attractive because it can help identify small molecule candidate compounds that have a high probability of proceeding to clinical use. Most FDA approved, first-in-class small molecules were identified from phenotypic screening. However, phenotypic screening using rodent models is labor intensive, low-throughput, and very expensive. As a novel alternative for small molecule screening, we have been developing gene expression disease profiles, termed the Transcriptional Disease Signature (TDS), as readout of small molecule screens for therapeutic molecules. In this concept, compounds that can reverse, or otherwise affect known disease-associated gene expression patterns in whole animals may be rapidly identified for more detailed downstream direct testing of their efficacy and mode of action. To establish proof of concept for this screening strategy, we employed a transgenic strain of a small aquarium fish, medaka (Oryzias latipes), that overexpresses the malignant melanoma driver gene xmrk, a mutant egfr gene, that is driven by a pigment cell-specific mitf promoter. In this model, melanoma develops with 100% penetrance. Using the transgenic medaka malignant melanoma model, we established a screening system that employs the NanoString nCounter platform to quantify gene expression within custom sets of TDS gene targets that we had previously shown to exhibit differential transcription among xmrk-transgenic and wild-type medaka. Compound-modulated gene expression was identified using an internet-accessible custom-built data processing pipeline. The effect of a given drug on the entire TDS profile was estimated by comparing compound-modulated genes in the TDS using an activation Z-score and Kolmogorov-Smirnov statistics. TDS gene probes were designed that target common signaling pathways that include proliferation, development, toxicity, immune function, metabolism and detoxification. These pathways may be utilized to evaluate candidate compounds for potential favorable, or unfavorable, effects on melanoma-associated gene expression. Here we present the logistics of using medaka to screen compounds, as well as, the development of a user-friendly NanoString data analysis pipeline to support feasibility of this novel TDS drug-screening strategy.
KW  - bioinformatics
KW  - phenotypic screening
Y1  - 2019
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-237322
VL  - 9
ER  - 
TY  - JOUR
A1  - Kraus, Michael
A1  - Grimm, Clemens
A1  - Seibel, Jürgen
T1  - Reversibility of a Point Mutation Induced Domain Shift: Expanding the Conformational Space of a Sucrose Phosphorylase
JF  - Scientific Reports
N2  - Despite their popularity as enzyme engineering targets structural information about Sucrose Phosphorylases remains scarce. We recently clarified that the Q345F variant of Bifidobacterium adolescentis Sucrose Phosphorylase is able to accept large polyphenolic substrates like resveratrol via a domain shift. Here we present a crystal structure of this variant in a conformation suitable for the accommodation of the donor substrate sucrose in excellent agreement with the wild type structure. Remarkably, this conformation does not feature the previously observed domain shift which is therefore reversible and part of a dynamic process rather than a static phenomenon. This crystallographic snapshot completes our understanding of the catalytic cycle of this useful variant and will allow for a more rational design of further generations of Sucrose Phosphorylase variants.
KW  - biocatalysis
KW  - X-ray crystallography
Y1  - 2018
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-224845
VL  - 8
ER  - 
TY  - JOUR
A1  - Kraus, Amelie J.
A1  - Brink, Benedikt G.
A1  - Siegel, T. Nicolai
T1  - Efficient and specific oligo-based depletion of rRNA
JF  - Scientific Reports
N2  - In most organisms, ribosomal RNA (rRNA) contributes to >85% of total RNA. Thus, to obtain useful information from RNA-sequencing (RNA-seq) analyses at reasonable sequencing depth, typically, mature polyadenylated transcripts are enriched or rRNA molecules are depleted. Targeted depletion of rRNA is particularly useful when studying transcripts lacking a poly(A) tail, such as some non-coding RNAs (ncRNAs), most bacterial RNAs and partially degraded or immature transcripts. While several commercially available kits allow effective rRNA depletion, their efficiency relies on a high degree of sequence homology between oligonucleotide probes and the target RNA. This restricts the use of such kits to a limited number of organisms with conserved rRNA sequences. In this study we describe the use of biotinylated oligos and streptavidin-coated paramagnetic beads for the efficient and specific depletion of trypanosomal rRNA. Our approach reduces the levels of the most abundant rRNA transcripts to less than 5% with minimal off-target effects. By adjusting the sequence of the oligonucleotide probes, our approach can be used to deplete rRNAs or other abundant transcripts independent of species. Thus, our protocol provides a useful alternative for rRNA removal where enrichment of polyadenylated transcripts is not an option and commercial kits for rRNA are not available.
KW  - parasite biology
KW  - RNA sequencing
KW  - transcriptomics
Y1  - 2019
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-224829
VL  - 9
ER  - 
TY  - JOUR
A1  - Kim, Bo-Mi
A1  - Amores, Angel
A1  - Kang, Seunghyun
A1  - Ahn, Do-Hwan
A1  - Kim, Jin-Hyoung
A1  - Kim, Il-Chan
A1  - Lee, Jun Hyuck
A1  - Lee, Sung Gu
A1  - Lee, Hyoungseok
A1  - Lee, Jungeun
A1  - Kim, Han-Woo
A1  - Desvignes, Thomas
A1  - Batzel, Peter
A1  - Sydes, Jason
A1  - Titus, Tom
A1  - Wilson, Catherine A.
A1  - Catchen, Julian M.
A1  - Warren, Wesley C.
A1  - Schartl, Manfred
A1  - Detrich, H. William III
A1  - Postlethwait, John H.
A1  - Park, Hyun
T1  - Antarctic blackfin icefish genome reveals adaptations to extreme environments
JF  - Nature Ecology & Evolution
N2  - Icefishes (suborder Notothenioidei; family Channichthyidae) are the only vertebrates that lack functional haemoglobin genes and red blood cells. Here, we report a high-quality genome assembly and linkage map for the Antarctic blackfin icefish Chaenocephalus aceratus, highlighting evolved genomic features for its unique physiology. Phylogenomic analysis revealed that Antarctic fish of the teleost suborder Notothenioidei, including icefishes, diverged from the stickleback lineage about 77 million years ago and subsequently evolved cold-adapted phenotypes as the Southern Ocean cooled to sub-zero temperatures. Our results show that genes involved in protection from ice damage, including genes encoding antifreeze glycoprotein and zona pellucida proteins, are highly expanded in the icefish genome. Furthermore, genes that encode enzymes that help to control cellular redox state, including members of the sod3 and nqo1 gene families, are expanded, probably as evolutionary adaptations to the relatively high concentration of oxygen dissolved in cold Antarctic waters. In contrast, some crucial regulators of circadian homeostasis (cry and per genes) are absent from the icefish genome, suggesting compromised control of biological rhythms in the polar light environment. The availability of the icefish genome sequence will accelerate our understanding of adaptation to extreme Antarctic environments.
KW  - animal physiology
KW  - evolutionary genetics
KW  - genomics
KW  - ichthyology
Y1  - 2019
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-325811
VL  - 3
ER  - 
TY  - JOUR
A1  - Hines, Rochelle M.
A1  - Maric, Hans Michael
A1  - Hines, Dustin J.
A1  - Modgil, Amit
A1  - Panzanelli, Patrizia
A1  - Nakamura, Yasuko
A1  - Nathanson, Anna J.
A1  - Cross, Alan
A1  - Deeb, Tarek
A1  - Brandon, Nicholas J.
A1  - Davies, Paul
A1  - Fritschy, Jean-Marc
A1  - Schindelin, Hermann
A1  - Moss, Stephen J.
T1  - Developmental seizures and mortality result from reducing GABAA receptor α2-subunit interaction with collybistin
JF  - Nature Communications
N2  - Fast inhibitory synaptic transmission is mediated by γ-aminobutyric acid type A receptors (GABAARs) that are enriched at functionally diverse synapses via mechanisms that remain unclear. Using isothermal titration calorimetry and complementary methods we demonstrate an exclusive low micromolar binding of collybistin to the α2-subunit of GABAARs. To explore the biological relevance of collybistin-α2-subunit selectivity, we generate mice with a mutation in the α2-subunit-collybistin binding region (Gabra2-1). The mutation results in loss of a distinct subset of inhibitory synapses and decreased amplitude of inhibitory synaptic currents. Gabra2–1 mice have a striking phenotype characterized by increased susceptibility to seizures and early mortality. Surviving Gabra2-1 mice show anxiety and elevations in electroencephalogram δ power, which are ameliorated by treatment with the α2/α3-selective positive modulator, AZD7325. Taken together, our results demonstrate an α2-subunit selective binding of collybistin, which plays a key role in patterned brain activity, particularly during development.
KW  - cellular neuroscience
KW  - ion channels in the nervous system
KW  - neurotransmitters
KW  - synaptic development
Y1  - 2018
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-320719
VL  - 9
ER  - 
TY  - JOUR
A1  - Hennrich, Marco L.
A1  - Romanov, Natalie
A1  - Horn, Patrick
A1  - Jaeger, Samira
A1  - Eckstein, Volker
A1  - Steeples, Violetta
A1  - Ye, Fei
A1  - Ding, Ximing
A1  - Poisa-Beiro, Laura
A1  - Mang, Ching Lai
A1  - Lang, Benjamin
A1  - Boultwood, Jacqueline
A1  - Luft, Thomas
A1  - Zaugg, Judith B.
A1  - Pellagatti, Andrea
A1  - Bork, Peer
A1  - Aloy, Patrick
A1  - Gavin, Anne-Claude
A1  - Ho, Anthony D.
T1  - Cell-specific proteome analyses of human bone marrow reveal molecular features of age-dependent functional decline
JF  - Nature Communications
N2  - Diminishing potential to replace damaged tissues is a hallmark for ageing of somatic stem cells, but the mechanisms remain elusive. Here, we present proteome-wide atlases of age-associated alterations in human haematopoietic stem and progenitor cells (HPCs) and five other cell populations that constitute the bone marrow niche. For each, the abundance of a large fraction of the ~12,000 proteins identified is assessed in 59 human subjects from different ages. As the HPCs become older, pathways in central carbon metabolism exhibit features reminiscent of the Warburg effect, where glycolytic intermediates are rerouted towards anabolism. Simultaneously, altered abundance of early regulators of HPC differentiation reveals a reduced functionality and a bias towards myeloid differentiation. Ageing causes alterations in the bone marrow niche too, and diminishes the functionality of the pathways involved in HPC homing. The data represent a valuable resource for further analyses, and for validation of knowledge gained from animal models.
KW  - ageing
KW  - haematopoietic stem cells
KW  - mesenchymal stem cells
Y1  - 2018
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-319877
VL  - 9
ER  - 
TY  - JOUR
A1  - Heil, Hannah S.
A1  - Schreiber, Benjamin
A1  - Götz, Ralph
A1  - Emmerling, Monika
A1  - Dabauvalle, Marie-Christine
A1  - Krohne, Georg
A1  - Höfling, Sven
A1  - Kamp, Martin
A1  - Sauer, Markus
A1  - Heinze, Katrin G.
T1  - Sharpening emitter localization in front of a tuned mirror
JF  - Light: Science & Applications
N2  - Single-molecule localization microscopy (SMLM) aims for maximized precision and a high signal-to-noise ratio1. Both features can be provided by placing the emitter in front of a metal-dielectric nanocoating that acts as a tuned mirror2,3,4. Here, we demonstrate that a higher photon yield at a lower background on biocompatible metal-dielectric nanocoatings substantially improves SMLM performance and increases the localization precision by up to a factor of two. The resolution improvement relies solely on easy-to-fabricate nanocoatings on standard glass coverslips and is spectrally and spatially tunable by the layer design and wavelength, as experimentally demonstrated for dual-color SMLM in cells.
KW  - imaging and sensing
KW  - super-resolution microscopy
Y1  - 2018
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-228080
VL  - 7
ER  - 
TY  - THES
A1  - Hahn, Sarah
T1  - Investigating non-canonical, 5' UTR-dependent translation of MYC and its impact on colorectal cancer development
T1  - Untersuchung der nicht-kanonischen, 5' UTR-abhängigen Translation von MYC und ihres Einflusses auf die Entwicklung von Darmkrebs
N2  - Colorectal cancer (CRC) is the second most common tumour disease in Germany, with the sequential accumulation of certain mutations playing a decisive role in the transition from adenoma to carcinoma. In particular, deregulation of the Wnt signalling pathway and the associated deregulated expression of the MYC oncoprotein play a crucial role. Targeting MYC thus represents an important therapeutic approach in the treatment of tumours. Since direct inhibition of MYC is challenging, various approaches have been pursued to date to target MYC indirectly. The MYC 5' UTR contains an internal ribosomal entry site (IRES), which has a particular role in the initiation of MYC translation, especially in multiple myeloma. As basis for this work, it was hypothesised on the basis of previous data that translation of MYC potentially occurs via its IRES in CRC as well. Based on this, two IRES inhibitors were tested for their potential to regulate MYC expression in CRC cells. In addition, alternative, 5’ UTR-dependent translation of MYC and interacting factors were investigated. EIF3D was identified as a MYC 5' UTR binding protein which has the potential to regulate MYC expression in CRC. The results of this work suggest that there is a link between eIF3D and MYC expression/translation, rendering eIF3D a potential therapeutic target for MYC-driven CRCs.
N2  - Das kolorektale Karzinom (KRK) ist die zweithäufigste Tumorerkrankung in Deutschland, wobei die sequenzielle Akkumulation bestimmter Mutationen eine entscheidende Rolle beim Übergang vom Adenom zum Karzinom spielt. Insbesondere die Deregulation des Wnt-Signalweges und die damit verbundene deregulierte Expression des MYC-Onkoproteins spielen eine entscheidende Rolle. MYC ist ein zentraler Vermittler von Zellfunktionen und reguliert als Transkriptionsfaktor die Expression fast aller Gene sowie verschiedener RNA-Spezies. Selbst kleine Veränderungen der zellulären MYC-Konzentration können das Proliferationsverhalten beeinflussen und die Entstehung und das Fortschreiten von Tumoren fördern. Die gezielte Beeinflussung von MYC stellt daher einen wichtigen therapeutischen Ansatz für die Behandlung von Tumoren dar.  Da eine direkte Hemmung von MYC aufgrund seiner Struktur herausfordernd ist, wurden bisher verschiedene Ansätze verfolgt, um MYC indirekt zu beeinflussen, etwa über seinen Interaktionspartner MAX oder auf Ebene der Stabilität, Transkription oder Translation. In unserer eigenen Forschungsgruppe lag der Schwerpunkt in den letzten Jahren speziell auf der Translation von MYC im KRK. Es konnte gezeigt werden, dass die Hemmung der kanonischen cap-abhängigen Translation nicht wie erwartet zu einer Verringerung der zellulären MYC-Level führt, was auf einen alternativen Mechanismus der MYC-Translation hindeutet, der unabhängig vom eIF4F-Komplex abläuft. Die 5'-UTR von MYC enthält eine interne ribosomale Eintrittsstelle (IRES), die eine besondere Rolle bei der Initiierung der MYC-Translation spielt, insbesondere im Multiplen Myelom. Als Grundlage für diese Arbeit wurde daher die Hypothese aufgestellt, dass die Translation von MYC im KRK möglicherweise ebenfalls über die IRES erfolgt. Auf dieser Grundlage wurden zunächst zwei publizierte IRES-Inhibitoren auf ihr Potenzial zur Regulierung der MYC-Expression in KRK-Zellen getestet. J007-IRES hatte keine Auswirkungen auf die MYC-Proteinmenge, und Cymarin scheint weitaus globalere Auswirkungen zu haben, die nicht ausschließlich auf die Verringerung der MYC-Proteinmenge zurückzuführen sind. Daher wurde weiter untersucht, inwieweit die alternative Translation von MYC generell von der 5'-UTR und damit interagierenden Faktoren abhängig ist. EIF3D wurde als MYC-5'-UTR-Bindungsprotein identifiziert, dessen Knockdown zu reduzierten MYC-Leveln, einem Proliferationsdefizit sowie einer Verringerung der globalen Proteinsynthese in KRK-Zellen führte. Darüber hinaus führte die Depletion von EIF3D zu ähnlichen Veränderungen im zellulären Genexpressionsmuster wie die Depletion von MYC, wobei viele tumorassoziierte Signalwege betroffen waren. Mittels eCLIP-seq wurde die Bindung von eIF3D an die MYC mRNA nachgewiesen, der genaue Mechanismus einer möglicherweise durch eIF3D vermittelten Translation von MYC muss jedoch weiter untersucht werden. Die Ergebnisse dieser Arbeit deuten darauf hin, dass eine Verbindung zwischen eIF3D und der MYC-Expression/Translation besteht, wodurch eIF3D zu einem potenziellen therapeutischen Ziel für MYC-getriebene KRKs wird.
KW  - Myc
KW  - Translation <Genetik>
KW  - Colorectal cancer
KW  - 5' UTR
Y1  - 2024
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-364202
ER  -