Human-Robot Gym: Benchmarking Reinforcement Learning in Human-Robot Collaboration

Deep reinforcement learning (RL) has shown promising results in robot motion planning with first attempts in human-robot collaboration (HRC). However, a fair comparison of RL approaches in HRC under the constraint of guaranteed safety is yet to be made. We, therefore, present human-robot gym, a benchmark for safe RL in HRC. Our benchmark provides eight challenging, realistic HRC tasks in a modular simulation framework. Most importantly, human-robot gym includes a safety shield that provably guarantees human safety. We are, thereby, the first to provide a benchmark to train RL agents that adhere to the safety specifications of real-world HRC. This bridges a critical gap between theoretic RL research and its real-world deployment. Our evaluation of six environments led to three key results: (a) the diverse nature of the tasks offered by human-robot gym creates a challenging benchmark for state-of-the-art RL methods, (b) incorporating expert knowledge in the RL training in the form of an action-based reward can outperform the expert, and (c) our agents negligibly overfit to training data

Functional Genomics of secretory Proteins and Host-Pathogen-Interactions of the human pathogen Listeria monocytogenes

In dieser Arbeit wurden zwei für das Humanpathogen L. monocytogenes wichtige Subproteome sowie die Wirtszellantwort auf Invasion dieses Pathogens mit Methoden der Funktionellen Genomanalyse untersucht. Mittels LC-MS/MS gelang erstmals, die kovalent-gebundenen Zellwandproteine von L. monocytogenes zu untersuchen und 6 LPXTG-Proteine wurden identifiziert. Weiterhin konnte gezeigt werden, dass diese für die Pathogenität von Listeria monocytogenes wichtige Proteinklasse über das Enzym Sortase A an das Peptidoglycan verknüpft wird. Durch 2D-Gelelektrophorese und LC-MS/MS wurden 105 sekretorische Proteine identifiziert, darunter alle primären Virulenzfaktoren. Durch einen Vergleich mit einer prfA-Deletionsmutante konnte die PrfA-abhängige Expression der primären Virulenzfaktoren bestätigt werden und 13 weitere putativ PrfA-regulierte Proteine identifiziert werden. Durch einen Vergleich der Überstandsproteine der apathogenen Art L. innocua mit L. monocytogenes wurden neben den 8 primären Virulenzfaktoren 8 weitere Proteine identifiziert, die ebenfalls keine orthologen Gene in der apathogenen Art L. innocua besitzen. Bei Proteom- und ersten Transkriptionsuntersuchungen von mit L. monocytogenes infizierten HeLa-Zellen wurden mehrere Hinweise gefunden, dass während der listeriellen Invasion anti-apoptotische Signalwege in den Wirtszellen induziert sein könnten. Weiterhin konnte gezeigt werden, dass das Mikrotubuli-destabilisierende Protein Stathmin durch Infektion mit Listerien spezifisch phosphoryliert wird. Durch die Präsentation von rekombinantem InlB und des natürlichen Liganden von Met, HGF, konnte bestätigt werden, dass die Aktivierung von Met dafür verantwortlich ist.In this work two subproteomes of the human pathogen L. monocytogenes and the host-cell response to listerial invasion were analysed by Functional Genomics. By LC-MS/MS the cell wall proteins of L. monocytogenes covalently bound to the peptidoglycan were analysed and 6 LPXTG-proteins identified. Furthermore was shown that the enzyme Sortase A attaches these proteins that are important for listerial pathogenicity to the cell surface. By 2D gel electrophoresis and LC-MS/MS 105 listerial secretory proteins were identified, among these all primary virulence factors. By comparison with a prfA-deletion mutant the PrfA-dependent expression of all primary virulence factors and further 13 putatively PrfA-regulated proteins could be identified. By comparison with the culture supernatant proteins of the apathogenic species L. innocua the 8 primary virulence factors and 8 further proteins without any orthologous gene in the apathogenic species were identified. By proteomics and preliminary trancriptomics analysis of eukaryotic cells infected with L. monocytogenes evidence for the induction of anti-apoptotic signalling in host-cells by listerial invasion was found. Furthermore was shown that the microtubule-destabilising protein stathmin is specifically phosphorylated during listerial invasion. By presentation of recombinant InlB and the natural ligand of Met, HGF, was shown, that the phosphorylation of stathmin is down-stream of activated Met

The MALDI TOF E2/E3 ligase assay as universal tool for drug discovery in the ubiquitin pathway

AbstractIn many diseases, components of the ubiquitin system - such as E2/E3 ligases and deubiquitylases - are dysregulated. The ubiquitin system has therefore become an emergent target for the treatment of a number of diseases, including cancer, neurodegeneration and autoimmunity. Despite of the efforts in this field, primary screenings of compound libraries to individuate new potential therapeutic molecules targeting the ubiquitin pathway have been strongly limited by the lack of robust and fast high-throughput assays. Here we report the first label-free high-throughput screening (HTS) assay for ubiquitin E2 conjugating enzymes and E3 ligases based on Matrix-Assisted Laser Desorption/Ionization Time-Of-Flight (MALDI TOF) mass spectrometry. The MALDI TOF E2/E3 assay allows us to test E2 conjugating enzymes and E3 ligases for their ubiquitin transfer activity, to identify E2/E3 active pairs, inhibitor potency and specificity and to screen compound librariesin vitrowithout synthesis of chemical or fluorescent probes. We demonstrate that the MALDI TOF E2/E3 assay is a universal tool for drug discovery screening in the ubiquitin pathway as it is suitable for working with all E3 ligase families and requires a reduced amount of reagents, compared to standard biochemical assays.</jats:p

Inhibiting Multiple Deubiquitinases to Reduce Androgen Receptor Expression in Prostate Cancer Cells

Prostate cancer (PCa), a leading cause of cancer-related death in men, becomes resistant to androgen deprivation therapy by inducing androgen receptor (AR) activity, which is known as castration-resistant PCa (CRPC). Enzalutamide is an approved drug that inhibits AR activity and increases overall survival. However, resistance to enzalutamide develops rapidly often by increasing AR activity, suggesting that new therapies are required for CRPC. We investigated whether betulinic acid (BA), a small molecule from plants that inhibits multiple deubiquitinases (DUBs), reduces AR, and selectively kills PCa cells, can provide an adjuvant strategy for CRPC. Our data indicated that BA reduced AR protein stability and mRNA expression, making it an attractive agent for CRPC. BA decreased AR mRNA possibly by inhibiting a histone 2A DUB thereby increasing ubiquitinated histone 2A, a transcriptional repressor. We identified multiple and specific DUBs inhibited by BA either in PCa cells or using recombinant DUBs. Similar results were obtained using another multi-DUB inhibitor WP1130, suggesting that these DUB inhibitors can decrease AR expression and increase PCa-specific death. Our results also suggest that combining multi-DUB inhibitors BA or WP1130 with enzalutamide may provide a novel strategy for CRPC by further decreasing AR expression and increasing apoptotic cell death.</p

Unanchored tri-NEDD8 inhibits PARP-1 to protect from oxidative stress-induced cell death

NEDD8 is a ubiquitin‐like protein that activates cullin‐RING E3 ubiquitin ligases (CRLs). Here, we identify a novel role for NEDD8 in regulating the activity of poly(ADP‐ribose) polymerase 1 (PARP‐1) in response to oxidative stress. We show that treatment of cells with H2O2 results in the accumulation of NEDD8 chains, likely by directly inhibiting the deneddylase NEDP1. One chain type, an unanchored NEDD8 trimer, specifically bound to the second zinc finger domain of PARP‐1 and attenuated its activation. In cells in which Nedp1 is deleted, large amounts of tri‐NEDD8 constitutively form, resulting in inhibition of PARP‐1 and protection from PARP‐1‐dependent cell death. Surprisingly, these NEDD8 trimers are additionally acetylated, as shown by mass spectrometry analysis, and their binding to PARP‐1 is reduced by the overexpression of histone de‐acetylases, which rescues PARP‐1 activation. Our data suggest that trimeric, acetylated NEDD8 attenuates PARP‐1 activation after oxidative stress, likely to delay the initiation of PARP‐1‐dependent cell death

Thermal proteome profiling of breast cancer cells reveals proteasomal activation by CDK4/6 inhibitor palbociclib

Palbociclib is a CDK4/6 inhibitor approved for metastatic estrogen receptor-positive breast cancer. In addition to G1 cell cycle arrest, palbociclib treatment results in cell senescence, a phenotype that is not readily explained by CDK4/6 inhibition. In order to identify a molecular mechanism responsible for palbociclib-induced senescence, we performed thermal proteome profiling of MCF7 breast cancer cells. In addition to affecting known CDK4/6 targets, palbociclib induces a thermal stabilization of the 20S proteasome, despite not directly binding to it. We further show that palbociclib treatment increases proteasome activity independently of the ubiquitin pathway. This leads to cellular senescence, which can be counteracted by proteasome inhibitors. Palbociclib-induced proteasome activation and senescence is mediated by reduced proteasomal association of ECM29. Loss of ECM29 activates the proteasome, blocks cell proliferation, and induces a senescence-like phenotype. Finally, we find that ECM29 mRNA levels are predictive of relapse-free survival in breast cancer patients treated with endocrine therapy. In conclusion, thermal proteome profiling identifies the proteasome and ECM29 protein as mediators of palbociclib activity in breast cancer cells

Is the proteomic composition of the salivary pellicle dependent on the substrate material?

Purpose: The use of dental restorative materials is a routine task in clinical dentistry. Upon exposure to the oral cavity, continuous adsorption of salivary proteins and other macromolecules to all surfaces occurs, representing the first step in dental biofilm formation. Different physico-chemical properties of substrate materials potentially influence the composition of the initial biofilm, termed pellicle. This study aimed at characterizing and comparing the individual proteomic composition of the 3-min pellicle formed on bovine enamel and six restorative materials. Experimental Design: After chemical elution, pellicle proteins were identified by nano-LC-HR-MS/MS. Proteomic profiles were analyzed in terms of molecular weights, isoelectric points, molecular functions and compared to saliva to reveal substrate material-specific adsorption patterns. Results: A total of 1348 different pellicle proteins were identified, with 187–686 proteins in individual 3-min pellicles. Unexpectedly, this yielded quite similar distribution patterns independent of the substrate materials. Furthermore, overall similar fold changes were obtained for the major part of commonly enriched or depleted proteins in the pellicles. Conclusions and Clinical Relevance: The current results point to a minor role of the substrate material on the proteomic composition of the 3-min pellicle and represent core data for understanding the complex surface interactions in the oral cavity

Killing with proficiency:integrated post-translational regulation of an offensive Type VI secretion system

<div><p>The Type VI secretion system (T6SS) is widely used by bacterial pathogens as an effective weapon against bacterial competitors and is also deployed against host eukaryotic cells in some cases. It is a contractile nanomachine which delivers toxic effector proteins directly into target cells by dynamic cycles of assembly and firing. Bacterial cells adopt distinct post-translational regulatory strategies for deployment of the T6SS. ‘Defensive’ T6SSs assemble and fire in response to incoming attacks from aggressive neighbouring cells, and can utilise the Threonine Protein Phosphorylation (TPP) regulatory pathway to achieve this control. However, many T6SSs are ‘offensive’, firing at all-comers without the need for incoming attack or other cell contact-dependent signal. Post-translational control of the offensive mode has been less well defined but can utilise components of the same TPP pathway. Here, we used the anti-bacterial T6SS of <i>Serratia marcescens</i> to elucidate post-translational regulation of offensive T6SS deployment, using single-cell microscopy and genetic analyses. We show that the integration of the TPP pathway with the negative regulator TagF to control core T6SS machine assembly is conserved between offensive and defensive T6SSs. Signal-dependent PpkA-mediated phosphorylation of Fha is required to overcome inhibition of membrane complex assembly by TagF, whilst PppA-mediated dephosphorylation promotes spatial reorientation and efficient killing. In contrast, the upstream input of the TPP pathway defines regulatory strategy, with a new periplasmic regulator, RtkS, shown to interact with the PpkA kinase in <i>S</i>. <i>marcescens</i>. We propose a model whereby the opposing actions of the TPP pathway and TagF impose a delay on T6SS re-assembly after firing, providing an opportunity for spatial re-orientation of the T6SS in order to maximise the efficiency of competitor cell targeting. Our findings provide a better understanding of how bacterial cells deploy competitive weapons effectively, with implications for the structure and dynamics of varied polymicrobial communities.</p></div