Einfluss von MTA auf die Differenzierung und Reifung humaner Monozyten zu dendritischen Zellen in vitro

Durch Modulation ihres Metabolismus sind maligne entartete Zellen häufig imstande, sich einen Überlebensvorteil gegenüber dem umgebenden Gewebe zu verschaffen. Beispielsweise zeigen Tumorzellen Veränderungen in der Polyaminsynthese: In vielen Tumoren lässt sich eine Defizienz der Methylthioadenosin-Phosphorylase nachweisen, wodurch es zu einer extrazellulären Akkumulation von 5’-Deoxy-5’-Methythioadenosin (MTA) kommt. Da bereits immunmodulatorische Effekte von MTA bekannt sind, wurde in der vorliegenden Arbeit die Wirkung von MTA auf die Reifung und Funktion humaner dendritischer Zellen (DCs) analysiert. Es konnte gezeigt werden, dass MTA einen hemmenden Einfluss auf die Antigenexpression wichtiger Oberflächenmarker auf DCs hat. Neben einer Modulation der Reifung kam es in Gegenwart von 150 µM MTA auch zu einer Suppression der IL-12-Produktion der DCs. In der gemischten Lymphozytenreaktion (MLR) waren zuvor mit MTA inkubierte DCs in ihrer Fähigkeit, die Proliferation von allogenen T-Zellen zu induzieren, eingeschränkt. Auch bei Zugabe von MTA in die MLR kam es zu einer reduzierten Aktivierung der T-Zellantwort. In einem weiteren Ansatz konnte gezeigt werden, dass MTA einen hemmenden Effekt auf die Proliferation polyklonal stimulierter T-Zellen hat und somit DCs und T-Zellen beeinflusst. Des Weiteren wurde eine Herunterregulation von wichtigen Komponenten der Antigenprozessierungsmaschinerie wie TAP1, TAP2 und LMP2 in DCs auf mRNA-Ebene gezeigt, was bei einigen Komponenten auch auf Proteinebene detektierbar war. Insgesamt zeigte sich bei den DCs eine Hemmung ihrer Reifung und eine Schwächung ihrer Funktion als antigenpräsentierenden Zellen durch MTA. Aufgrund der beobachteten Hemmung der Reifung von DCs und der Schwächung ihrer Funktion als antigenpräsentierenden Zellen durch MTA, stellt die Sekretion von MTA durch Tumorzellen einen immune escape-Mechanismus dar, die dem Tumor durch Beeinträchtigung der Immunantwort einen Wachstumsvorteil verschafft. Die weitere Erforschung des Tumormetabolismus und seine Auswirkung auf das Immunsystem ist folglich wesentlich, um eine erfolgreiche Antitumortherapie zu konzipieren. In zukünftigen Experimenten sollten hierfür auch mögliche Wirkmechanismen, durch die MTA DCs und andere Immunzellen moduliert, untersucht werden

Behinderung und berufliche Rehabilitation in Deutschland und der Schweiz

Wie ist ein rechtlicher Behinderungsbegriff auszugestalten, welche Maßnahmen der beruflichen Rehabilitation werden seitens des Staates angeboten und wie werden Dritte in die Leistungserbringung eingebunden? Diesen Fragen geht die Autorin in einer rechtvergleichenden Untersuchung zwischen Deutschland und der Schweiz nach. In einer abschließenden Betrachtung wird der Bogen zwischen der Untersuchung positiven Rechts zur rechtstheoretischen Frage gespannt, welchen Wert ein Rechtsvergleich hat. Dass ein Rechtsvergleich ein Hilfsmittel zur Systematisierung eigenen Rechts sein kann, wird anhand konkret gewonnener Erkenntnisse aus dem Rechtsvergleich belegt

A Local Role for the Small Ribosomal Subunit Primary Binder rpS5 in Final 18S rRNA Processing in Yeast

In vivo depletion of the yeast small ribosomal subunit (SSU) protein S5 (rpS5) leads to nuclear degradation of nascent SSUs and to a perturbed global assembly state of the SSU head domain. Here, we report that rpS5 plays an additional local role at the head/platform interface in efficient SSU maturation. We find that yeast small ribosomal subunits which incorporated an rpS5 variant lacking the seven C-terminal amino acids have a largely assembled head domain and are exported to the cytoplasm. On the other hand, 3′ processing of 18S rRNA precursors is inhibited in these ribosomal particles, although they associate with the putative endonuclease Nob1p and other late acting 40S biogenesis factors. We suggest that the SSU head component rpS5 and platform components as rpS14 are crucial constituents of a highly defined spatial arrangement in the head – platform interface of nascent SSUs, which is required for efficient processing of the therein predicted SSU rRNA 3′ end. Positioning of rpS5 in nascent SSUs, including its relative orientation towards platform components in the head-platform cleft, will depend on the general assembly and folding state of the head domain. Therefore, the suggested model can explain 18S precursor rRNA 3′ processing phenotypes observed in many eukaryotic SSU head assembly mutants

The pathogenic exon 1 HTT protein is produced by incomplete splicing in Huntington’s disease patients

We have previously shown that exon 1 of the huntingtin gene does not always splice to exon 2 resulting in the production of a small polyadenylated mRNA (HTTexon1) that encodes the highly pathogenic exon 1 HTT protein. The level of this read-through product is proportional to CAG repeat length and is present in all knock-in mouse models of Huntington’s disease (HD) with CAG lengths of 50 and above and in the YAC128 and BACHD mouse models, both of which express a copy of the human HTT gene. We have now developed specific protocols for the quantitative analysis of the transcript levels of HTTexon1 in human tissue and applied these to a series of fibroblast lines and post-mortem brain samples from individuals with either adult-onset or juvenile-onset HD. We found that the HTTexon1 mRNA is present in fibroblasts from juvenile HD patients and can also be readily detected in the sensory motor cortex, hippocampus and cerebellum of post-mortem brains from HD individuals, particularly in those with early onset disease. This finding will have important implications for strategies to lower mutant HTT levels in patients and the design of future therapeutics

Regulatory mechanisms of incomplete huntingtin mRNA splicing

Huntington's disease is caused by a CAG repeat expansion in exon 1 of the HTT gene. We have previously shown that exon 1 HTT does not always splice to exon 2 producing a small transcript (HTTexon1) that encodes the highly pathogenic exon 1 HTT protein. The mechanisms by which this incomplete splicing occurs are unknown. Here, we have generated a minigene system that recapitulates the CAG repeat-length dependence of HTTexon1 production, and has allowed us to define the regions of intron 1 necessary for incomplete splicing. We show that manipulation of the expression levels of the splicing factor SRSF6, predicted to bind CAG repeats, modulates this aberrant splicing event and also demonstrate that RNA polymerase II transcription speed regulates the levels of HTTexon1 production. Understanding the mechanisms by which this pathogenic exon 1 HTT is generated may provide the basis for the development of strategies to prevent its production

rRNA Maturation in Yeast Cells Depleted of Large Ribosomal Subunit Proteins

The structural constituents of the large eukaryotic ribosomal subunit are 3 ribosomal RNAs, namely the 25S, 5.8S and 5S rRNA and about 46 ribosomal proteins (r-proteins). They assemble and mature in a highly dynamic process that involves more than 150 proteins and 70 small RNAs. Ribosome biogenesis starts in the nucleolus, continues in the nucleoplasm and is completed after nucleo-cytoplasmic translocation of the subunits in the cytoplasm. In this work we created 26 yeast strains, each of which conditionally expresses one of the large ribosomal subunit (LSU) proteins. In vivo depletion of the analysed LSU r-proteins was lethal and led to destabilisation and degradation of the LSU and/or its precursors. Detailed steady state and metabolic pulse labelling analyses of rRNA precursors in these mutant strains showed that LSU r-proteins can be grouped according to their requirement for efficient progression of different steps of large ribosomal subunit maturation. Comparative analyses of the observed phenotypes and the nature of r-protein – rRNA interactions as predicted by current atomic LSU structure models led us to discuss working hypotheses on i) how individual r-proteins control the productive processing of the major 5′ end of 5.8S rRNA precursors by exonucleases Rat1p and Xrn1p, and ii) the nature of structural characteristics of nascent LSUs that are required for cytoplasmic accumulation of nascent subunits but are nonessential for most of the nuclear LSU pre-rRNA processing events

Investigating the communicative function of breathing and non-breathing "silent" pauses

Cwiek A, Neueder S, Wagner P. Investigating the communicative function of breathing and non-breathing "silent" pauses. In: Draxler C, Kleber F, eds. Tagungsband der 12. Tagung Phonetik und Phonologie im deutschsprachigen Raum. München, Deutschland: Ludwig-Maximilians-Universität München; 2016: 27-29

PolyQ length-dependent metabolic alterations and DNA damage drive human astrocyte dysfunction in Huntington's disease

Huntington's Disease (HD) is a neurodegenerative disease caused by a polyglutamine (polyQ) expansion in the Huntingtin gene. Astrocyte dysfunction is known to contribute to HD pathology, however our understanding of the molecular pathways involved is limited. Transcriptomic analysis of patient-derived PSC (pluripotent stem cells) astrocyte lines revealed that astrocytes with similar polyQ lengths shared a large number of differentially expressed genes (DEGs). Notably, weighted correlation network analysis (WGCNA) modules from iPSC derived astrocytes showed significant overlap with WGCNA modules from two post-mortem HD cohorts. Further experiments revealed two key elements of astrocyte dysfunction. Firstly, expression of genes linked to astrocyte reactivity, as well as metabolic changes were polyQ length-dependent. Hypermetabolism was observed in shorter polyQ length astrocytes compared to controls, whereas metabolic activity and release of metabolites were significantly reduced in astrocytes with increasing polyQ lengths. Secondly, all HD astrocytes showed increased DNA damage, DNA damage response and upregulation of mismatch repair genes and proteins. Together our study shows for the first time polyQ-dependent phenotypes and functional changes in HD astrocytes providing evidence that increased DNA damage and DNA damage response could contribute to HD astrocyte dysfunction

Effects of context pre-exposure and delay until anxiety retrieval on generalization of contextual anxiety

Animal studies suggest that time delay between acquisition and retrieval of contextual anxiety increases generalization. Moreover, such generalization is prevented by preexposure to the context (CTX), presumably due to an improved representation of such context. We investigated whether preexposure and time-passing modulate generalization of contextual anxiety, in humans. On Day 1, 42 participants (preexposure group) explored two virtual offices, while 41 participants (no-preexposure group) explored a virtual stadium. On Day 2 (24 h later), all participants learned to associate one office (CTX+) with unpredictable unconditioned stimuli (USs), and another office (CTX2) with safety. On Day 3, either 24 h (recent test) or 2 wk (remote test) later, participants revisited CTX2 and CTX+ without USs, as well as a generalization context (G-CTX). Results revealed successfully conditioned anxiety and anxiety generalization for ratings (G-CTX was as aversive as CTX+ was), while safety generalization was found for startle responses (G-CTX elicited startle attenuation as CTX2 did). Time between learning and testing enhanced generalization as reflected by comparable startle responses to all three offices in the remote test. Contextual preexposure facilitated extinction of explicit conditioned anxiety assessed with ratings. These results suggest that memory trace of a context degrades with passage of time in humans like in animals and, consequently, anxiety generalization enhances. After context preexposure, high cognitive processes seem to be crucially involved in facilitating extinction (or safety) learning