C-Reactive Protein Causes Blood Pressure Drop in Rabbits and Induces Intracellular Calcium Signaling

Systemic diseases characterized by elevated levels of C-reactive protein (CRP), such as sepsis or systemic inflammatory response syndrome, are usually associated with hardly controllable haemodynamic instability. We therefore investigated whether CRP itself influences blood pressure and heart rate. Immediately after intravenous injection of purified human CRP (3.5 mg CRP/kg body weight) into anesthetized rabbits, blood pressure dropped critically in all animals, while control animals injected with bovine serum albumin showed no response. Heart rate did not change in either group. Approaching this impact on a cellular level, we investigated the effect of CRP in cell lines expressing adrenoceptors (CHO-α1A and DU-145). CRP caused a Ca(2+)signaling being dependent on the CRP dose. After complete activation of the adrenoceptors by agonists, CRP caused additional intracellular Ca(2+)mobilization. We assume that CRP interacts with hitherto unknown structures on the surface of vital cells and thus interferes with the desensitization of adrenoceptors

Monogenic variants in dystonia: an exome-wide sequencing study

Background Dystonia is a clinically and genetically heterogeneous condition that occurs in isolation (isolated dystonia), in combination with other movement disorders (combined dystonia), or in the context of multisymptomatic phenotypes (isolated or combined dystonia with other neurological involvement). However, our understanding of its aetiology is still incomplete. We aimed to elucidate the monogenic causes for the major clinical categories of dystonia. Methods For this exome-wide sequencing study, study participants were identified at 33 movement-disorder and neuropaediatric specialty centres in Austria, Czech Republic, France, Germany, Poland, Slovakia, and Switzerland. Each individual with dystonia was diagnosed in accordance with the dystonia consensus definition. Index cases were eligible for this study if they had no previous genetic diagnosis and no indication of an acquired cause of their illness. The second criterion was not applied to a subset of participants with a working clinical diagnosis of dystonic cerebral palsy. Genomic DNA was extracted from blood of participants and whole-exome sequenced. To find causative variants in known disorder-associated genes, all variants were filtered, and unreported variants were classified according to American College of Medical Genetics and Genomics guidelines. All considered variants were reviewed in expert round-table sessions to validate their clinical significance. Variants that survived filtering and interpretation procedures were defined as diagnostic variants. In the cases that went undiagnosed, candidate dystonia-causing genes were prioritised in a stepwise workflow. Findings We sequenced the exomes of 764 individuals with dystonia and 346 healthy parents who were recruited between June 1, 2015, and July 31, 2019. We identified causative or probable causative variants in 135 (19%) of 728 families, involving 78 distinct monogenic disorders. We observed a larger proportion of individuals with diagnostic variants in those with dystonia (either isolated or combined) with coexisting non-movement disorder-related neurological symptoms (100 [45%] of 222;excepting cases with evidence of perinatal brain injury) than in those with combined (19 [19%] of 98) or isolated (16 [4%] of 388) dystonia. Across all categories of dystonia, 104 (65%) of the 160 detected variants affected genes which are associated with neurodevelopmental disorders. We found diagnostic variants in 11 genes not previously linked to dystonia, and propose a predictive clinical score that could guide the implementation of exome sequencing in routine diagnostics. In cases without perinatal sentinel events, genomic alterations contributed substantively to the diagnosis of dystonic cerebral palsy. In 15 families, we delineated 12 candidate genes. These include IMPDH2, encoding a key purine biosynthetic enzyme, for which robust evidence existed for its involvement in a neurodevelopmental disorder with dystonia. We identified six variants in IMPDH2, collected from four independent cohorts, that were predicted to be deleterious de-novo variants and expected to result in deregulation of purine metabolism. Interpretation In this study, we have determined the role of monogenic variants across the range of dystonic disorders, providing guidance for the introduction of personalised care strategies and fostering follow-up pathophysiological explorations

Identification and characterization of ERG regulated signaling pathways in acute leukemia

Das ETS-related Gene ERG gehört zur Familie der ETS Transkriptionsfaktoren (TF) und übt Funktionen in der frühen zellulären Entwicklung aus und ist essentiell für die Aufrechterhaltung einer normalen Hämatopoese [49-51, 56]. Darüber hinaus kommen Fusionsgene zwischen ERG und verschiedenen Funktionspartnern bei malignen Erkrankungen wie der akuten myeloischen Leukämie (AML), dem Ewing Sarkom und Prostatakarzinomen vor [67, 76, 82]. Des Weiteren konnte eine hohe ERG-Expression als ungünstiger prognostischer Faktor in der AML und der akuten T-lymphoblastischen Leukämie (T-ALL) identifiziert werden [71, 72]. Trotz großer Fortschritte in den Therapieoptionen in den letzten Jahren gibt es immer noch Patienten mit AML oder T-ALL, deren Prognose auf Grund eines schlechten Therapieansprechens und hoher Rezidivraten schlecht ist. Eine solche Gruppe von Patienten sind jene mit hoher ERG-Expression, für die ein möglicher Therapieansatz die Inhibition ERG-regulierter Signalwege sein könnte. Allerdings sind bisher nur einige wenige Gene, deren Transkription durch ERG reguliert wird (Targetgene), bekannt, so dass das Ziel dieser Arbeit war, Targetgene des TFs ERG zu identifizieren, um somit mögliche Ansätze für neue Therapieoptionen der akuten Leukämien zu entwickeln. Daher wurden Chromatin-Immunopräzipitation on chip (ChIP-chip) Versuche an leukämischen Zelllinien (Jurkat und HL60) und primären Leukämieproben (5 AML, 1 T-ALL) durchgeführt. Nach statistischer Auswertung wurden die generierten Genlisten einer bioinformatischen Analyse unterzogen. Dabei zeigte sich eine signifikante Anreicherung des WNT/β-Catenin Signalwegs in der Zelllinie Jurkat sowie AML D und T-ALL (p<0,05). Des Weiteren konnte eine Anreicherung von Signalwegen wie PI3K/AKT, HMGB1, TNFR1, CNTF, p53 und Interferon Signalwege gezeigt werden. Ferner waren Prozesse der Zellmigration und –adhäsion sowie der zellulären Entwicklung und Regulation, Zellwachstum und Proliferation und Prozesse multizellulärer Organismen angereichert (p<0,05). Da diese Prozesse im Rahmen der Leukämogenese Veränderungen unterliegen, ist es denkbar, dass ERG durch die Beeinflussung dieser biologischen Funktionen ursächlich an der Leukämogenese beteiligt ist. Zur Validierung von Targetgenen im Jurkat ChIP- chip mittels quantitativer real-time Polymerasekettenreaktion (qrt-PCR) wurden 23 Gene mit Funktionen in der Hämatopoese, Onkogenese und WNT/β-Catenin Signalweg ausgewählt. Für 17 dieser Gene konnte eine Anreicherung bestätigt werden. Zur Validierung mittels qrt-PCR in den primären Leukämieproben wurden 5, in der Literatur beschriebene, mit ERG koregulierte Gene ausgewählt. Von diesen konnten WNT11 und DAAM1 in 5 bzw. allen 6 Proben durch eine mindestens 2-fache Anreicherung gegenüber der IgG-Kontrolle bestätigt werden. Die übrigen Gene zeigten lediglich eine Anreicherung in einer der Proben. Die Expression der validierten Gene wurde mittels ERG knock down und Überexpression untersucht. Dabei zeigte sich eine konstante und signifikante Verminderung der Expression des Gens WNT11 mit der Reduktion der ERG-Expression durch knock down und eine Induktion der WNT11-Expression durch ERG-Induktion. Dies spricht dafür, dass ERG als Transkriptionsaktivator von WNT11 fungiert. Weiterhin wurde die Proliferation ERG exprimierender Zellen mittels WST-Test bestimmt. Dabei zeigt sich ein Proliferationsvorteil der Zellen mit ERG-Induktion gegenüber Zellen ohne ERG-Expression bei der Behandlung mit dem Inhibitor der Glykogen-Synthase-Kinase 3β, BIO und dem Multikinaseinhibitor Sorafenib. Dies lässt vermuten, dass sich Zellen mit hoher ERG-Expression dem antiproliferativen Effekt dieser Substanzen entziehen, indem sie zur vermehrten Expression von Gegenspielern, wie z.B. WNT11, beitragen. Daraus ergibt sich, dass die Substanzen BIO und Sorafenib keine geeignete Therapieoption für akute Leukämien mit hoher ERG-Expression darstellen. Zusammenfassend zeigen diese Ergebnisse, dass ERG die Expression intrazellulärer Signalkaskaden wie WNT/β-Catenin, PI3K/AKT, HMGB1, TNFR1, CNTF, p53 und Interferon Signalwege reguliert und das Beispiel der Resistenz ERG exprimierender Zellen gegenüber BIO und Sorafenib verdeutlicht die Notwendigkeit der weiteren Exploration dieser Vorgänge, um gezieltere Therapien zu entwickeln.The ETS related gene ERG is a member of the ETS family of transcription factors (TF) and is associated with fundamental functions in cellular development. Furthermore, ERG has been implicated to be essential for the maintenance of a normal hematopoietic stem cell pool. Translocations involving the ERG locus result in fusion genes that can be found in acute myeloid leukemia (AML), Ewing sarcoma and prostate cancer. High ERG mRNA expression is an independent negative prognostic factor in AML and acute t-lymphoblastic leukemia (T-ALL). Despite great advances in the treatment of AML and T-ALL, there is still a great amount of patients with poor prognosis due to failure of first line therapy and early relapse. Hence, patients with high ERG expression might possibly profit from a therapeutic strategy aiming at the inhibition of ERG-regulated signaling pathways or ERG target genes. However, so far there are only very few ERG-regulated target genes identified. Thus, the main goal of this work was to identify ERG-regulated target genes as potential therapeutic targets in acute leukemia. Herein, chromatin- immunoprecipitation-on-chip (ChIP-chip) was applied on 8 samples (T-ALL cell line Jurkat, HL60 cell line as negative control and 6 primary leukemia samples – 5 AML, 1 T-ALL). Following statistical analysis, the enriched gene lists were analyzed using bioinformatics. Bioinformatic analysis revealed an enrichment of WNT/β-catenin, PI3K/AKT, TNFR1, interleukin, and p53 signaling pathways. Furthermore, fundamental biological functions enriched among ERG target genes included cellular migration and adhesion, cellular development, and proliferation. Potentially, these functions are influenced by ERG and altered in their activity during ERG-mediated leukemogenesis. In order to validate target genes, 23 genes with functions in hematopoiesis, oncogenesis, and WNT/β-catenin signaling were chosen for validation by quantitative real- time polymerase chain reaction (qrt-PCR). Seventeen were validated. The expression of these genes was investigated under the influence of the ERG expression. Thereby, the expression of WNT11 could be reduced by ERG knock down and induced by ERG overexpression. Thus, ERG seems to be a transcriptional activator of WNT11. Proliferation of ERG expressing cells has been measured by WST assay. ERG expressing cells show a proliferative advantage over cells without ERG expression when treated with the protein kinases BIO and sorafenib, suggesting an ERG-mediated resistance to the treatment with these substances. It is likely, that cells with high ERG expression escape the anti-proliferative effect of BIO and sorafenib by an ERG-mediated intrinsic resistance due to the synthesis of antagonizing proteins, like WNT11. In conclusion, this work shows, that ERG regulates the expression of genes associated with intracellular signaling cascades like WNT/β-catenin, PI3K/AKT, TNFR1, interleukin, and p53. The example of the ERG- mediated resistance to the treatment with BIO and sorafenib enhances the necessity of further exploration of these correlations in order to optimize therapeutic strategies

in Escherichia coli

for temperature-controlled gene expressio

Zielstrukturen in Familienunternehmen: Empirische Hinweise auf die Beziehung zwischen Unternehmens- und Familienzielen

In dieser Arbeit werden auf Basis einer Befragung von Eigentümern deutscher Unternehmen die Zielstrukturen in Familienunternehmen untersucht. Unsere Untersuchung zeigt, dass die Bedeutung, die Familien- und Unternehmenszielen beigemessen wird, von bestimmten Unternehmensmerkmalen abhängt. Ein starker Einfluss der Eigentümerfamilie begünstigt das Verfolgen von Familienzielen. In Übereinstimmung mit anderen Studien zeigen unsere Ergebnisse, dass die Existenz eines Beirates / Aufsichtsrates in negativem Zusammenhang zu Familienzielen steht. Größere Unternehmen zeigen hingegen eine stärkere Orientierung an Familienzielen

CLARIAH-DE work package 3: skills training and promotion of junior researchers

This poster summarizes the results of the CLARIAH-DE Work Package 3: Skills Training and Promotion of Junior Researchers. For a research field that is characterised by rapid technical development, CLARIAH-DE has to include the promotion of data literacy necessary for the efficient use of this digital research infrastructure as part of its objective. To develop, consolidate and refine a common programme in this area, work package 3 set itself the following sub goals: - Consolidation of the activities from the previous projects into a joint service - Cataloguing and reflecting on the methods and tools used in the research field, with the aim of identifying remaining gaps - Skills training of, individual support for and the promotion of junior researcher

Publisher Correction: An epigenetic gene silencing pathway selectively acting on transgenic DNA in the green alga Chlamydomonasis.

In the original version of this Article, the y-axis label of Fig. 1d was incorrectly given as “Relative H3Ac/H3 levels” and should read “Relative H4Ac/H3 levels”. In addition, the probe target in the left panel of Figure 1f was misspelled as “Guliver” and should read “Gulliver”. Furthermore, the last label of the diagram on the top of Fig. 2c was given incorrectly as “PF26” and should read “PF25”. Moreover, the label of the branch pointed by the arrow symbol in cluster II of the phylogenetic tree in Fig. 3c was incorrectly labelled as “CrSRTb” and the correct label is “CrSRTB”. These errors have been corrected in both the PDF and HTML versions of the Article

An epigenetic gene silencing pathway selectively acting on transgenic DNA in the green alga Chlamydomonas.

Silencing of exogenous DNA can make transgene expression very inefficient. Genetic screens in the model alga Chlamydomonas have demonstrated that transgene silencing can be overcome by mutations in unknown gene(s), thus producing algal strains that stably express foreign genes to high levels. Here, we show that the silencing mechanism specifically acts on transgenic DNA. Once a permissive chromatin structure has assembled, transgene expression can persist even in the absence of mutations disrupting the silencing pathway. We have identified the gene conferring the silencing and show it to encode a sirtuin-type histone deacetylase. Loss of gene function does not appreciably affect endogenous gene expression. Our data suggest that transgenic DNA is recognized and then quickly inactivated by the assembly of a repressive chromatin structure composed of deacetylated histones. We propose that this mechanism may have evolved to provide protection from potentially harmful types of environmental DNA