Technical Research Priorities for Big Data

To drive innovation and competitiveness, organisations need to foster the development and broad adoption of data technologies, value-adding use cases and sustainable business models. Enabling an effective data ecosystem requires overcoming several technical challenges associated with the cost and complexity of management, processing, analysis and utilisation of data. This chapter details a community-driven initiative to identify and characterise the key technical research priorities for research and development in data technologies. The chapter examines the systemic and structured methodology used to gather inputs from over 200 stakeholder organisations. The result of the process identified five key technical research priorities in the areas of data management, data processing, data analytics, data visualisation and user interactions, and data protection, together with 28 sub-level challenges. The process also highlighted the important role of data standardisation, data engineering and DevOps for Big Data

RNA Funktionskontrolle durch strukturelles Design

Das Hairpin-Ribozym-basierte System CRZ-2 wurde verwendet, um Selbst-Prozessierungsprodukte nach Ribozym-Reaktion zu untersuchen. Inter- und intramolekulare Ribozym-Reaktionen sollten mit CRZ-2 und dem entsprechenden linearen 83mer (l-83mer) durchgeführt und Oligomere und zyklisierte RNAs nachgewiesen werden. Über die Bildung der intermediären Spaltprodukte und des finalen Spaltproduktes konnte der Ablauf der Spalt-Kaskade komplett gezeigt werden. Dies war insbesondere durch vergleichende Verwendung von Test-Systemen im denaturierenden Polyacrylamid-Gel möglich, da eines der Systeme eine eindeutige Separation der Produkte im Gel zulässt. Weiter konnten die zwei erwarteten Spalt-Produkte (83mer und 92mer) über Sequenzierung ihrer komplementären DNAs nachgewiesen werden. Um zwischen zyklischen und linearen Reaktionsprodukten unterscheiden zu können, wurden folgende Methoden herangezogen: i) 2D-PAGE ii) exonukleolytischer Abbau von RNA in Lösung und im Gel, iii) Vergleich des Laufverhaltens eines inaktiven RNA-Monomers mit dem Reaktionsgemisch des l-83mer nach Ligationsreaktion, iv) AFM und v) Ferguson-Plot. Es zeigte sich, dass das CRZ-2 nach Ribozym-Reaktion ausschließlich lineare Produkte und Oligomere bis zum Trimer hervorbringt, während das l-83mer nach Ligationsreaktion auch einen Ring, das zyklische 83mer, hervorbringt. Das Wissen um die Art der Reaktionsprodukte von CRZ-2 und dem l-83mer ermöglichten es, diese beiden RNAs als Referenz-Systeme zu benutzen, um Hairpin-Ribozym-basierte Varianten zu untersuchen. Die bioinformatische Entwicklung neuer Varianten erfolgte in Kooperation mit der Arbeitsgruppe von Prof. Ivo Hofacker (Universität Wien). Dabei wurde ein wahrscheinlichkeitsbasierter Entwurf (probability based design, kurz: PBD) für RNA-Sekundärstrukturen mittels Programm Switch.pl aus dem Vienna RNA package 2.0 verwendet. Die für die katalytische Aktivität der Hairpin-Ribozym-Varianten essentiellen Basenfolgen in den Loops wurden beibehalten. Alle Test-Systeme waren bistabil, denn sie zeigten indirekt, das die für CRZ-2 übliche Spalt-Kaskade durchlaufen wurde, indem das jeweilige zyklische 83mer über 2D-PAGE nachgewiesen wurde. Wie erwartet, waren die Varianten insgesamt aktiver als das Referenzsystem CRZ-2 und sie unterschieden sich, wie bioinformatisch charakterisiert, in ihrem Zirkularisationsverhalten bezüglich der Monomere. Bioinformatisch unerwartet war das Zyklisierungsverhalten der Dimere. Ausschließlich bei Test-System 4 könnten gemäß 2D-PAGE und AFM Analyse unter anderem auch dimere RNA-Ring entstanden sein. PBD4 ist das System, bei welchem die geringste Dimer-Zyklisierungstendenz angenommen wurde. Eine erste Evaluierung der PBD-Methode ergibt somit, dass die Erwartungen für die bioinformatische Charakterisierung des Ablaufs der Spalt-Kaskade bis zur Monomerzyklisierung erfüllt wurden. Für die bioinformatisch nicht vorausgesagten experimentellen Ergebnisse, z.B. bei der Dimerzyklisierung, könnten verstärkt tertiäre Interaktionen relevant sein, die mit der PBD-Methode zur Sekundärstrukturvorhersage nicht berücksichtigt werden. Sequenz-Alignments der Test-Systeme zu CRZ-2 ließen Rückschlüsse auf die Funktionen einzelner Basen zu. Vier Basen traten zusätzlich zu den vorgegebenen Sequenzen auf. Diese befinden sich in einer Helix und könnten kritisch für das Zustandekommen dieser Helix als wichtiges Strukturelement im bistabilen Ribozym sein. Dieser Aspekt könnte in weiterführenden Arbeiten mit rationalem Design z.B. über Einfach- oder Doppelmutation weiter untersucht und die Auswirkungen auf die Selbst-Prozessierungsereignisse analysiert werden. Interessant sind auch die eingeführten Mutationen im superstabilen Tetraloop der Hairpin-Ribozym-Varianten. Im Sequenz-Alignment zeigte sich, dass sich PBD3 und 4 nur um zwei sich im Tetraloop befindlichen Basen unterscheiden (Positionen 1 und 3). Da sich die beiden Systeme bezüglich ihrer Oligomerisierungs- und Zyklisierungstendenz unterscheiden, sind diese beiden Positionen für die Funktionen essentiell.The hairpin ribozyme-based system CRZ-2 was used to investigate the self-processing products derived from ribozyme reaction. Inter- as well as intramolecular ribozyme reactions were performed with CRZ-2 and its linear 83mer (l-83mer), whereas oligomers and circularized RNAs were identified. Complete transition through the CRZ-2’s cleavage cascade was shown by the identification of intermediate and final cleavage products inter alia via Sanger sequencing of the corresponding complementary DNAs of the 83mer and the 92mer. Additionally, comparative polyacrylamide electrophoresis of test systems allowed to assign unambiguously the cleavage products of all systems in denatured gels. To distinguish between linear and circular products of ribozyme reactions several methods were combined: i) 2D-PAGE ii) assessment of exonucleolytic decay of RNA in solution and in gels, iii) comparison of the migration manner of an inactive RNA monomer with the reaction mixture of the l-83mer after ligation reaction iv) AFM and v) Ferguson-Plot. It became apparent that linear products such as oligomers up to trimers are exclusively derived from CRZ-2’s ribozyme reaction, while the l-83mer also forms RNA rings (the cyclic 83mer). The analyses of the reaction products of CRZ-2 and the l-83mer enabled their usage as reference systems for the investigation of hairpin ribozyme variants. The bioinformatic development of new CRZ-2 variants was performed in collaboration with the group of prof. Ivo Hofacker (University of Vienna) using a probability based design, short: PBD. The program Switch.pl from Vienna RNA package 2.0 was used to calculate the probability to form catalytically active secondary structures, resulting in hairpin ribozyme variants. The specific activity was ensured by keeping the essential bases of the loops while all other positions were freely exchangeable. All four test systems were bistable, displaying the cleavage cascade typical for CRZ-2. This was evaluated by identification of the cyclic 83mer by 2D-PAGE for all test systems. All CRZ-2 variants were more active than the reference system CRZ-2, and as bioinformatically proposed, they differ in their ability to form cyclic monomers. From a bioinformatic point of view the dimer circularization abilities of the test systems were unexpected. Test system four was the only variant that showed several RNA rings of different size, very likely also including cyclic dimers.PBD4 is the system which was meant to show the lowest ability to form cyclic dimers. A first evaluation of the probability based design reveals that bioinformatic expectations were fulfilled on a monomeric level from cleavage cascade to circularization of the 83mer. Bioinformatically non-expected experimental results may be caused by tertiary interactions that are not part of the PBD, dealing with predictions concerning secondary structures exclusively. Sequence alignments comparing test systems with CRZ-2 allowed for conclusions concerning the base functions. In addition to the essential bases which were not allowed to be changed in the PBD approach, four bases were found in all sequences. These four bases are present in helices, and may be essential for helix formation as an important structural motive in the bistable ribozyme. In future projects, this aspect can be further elucidated by single-or double mutations in the respective regions to assess their influence on self-processing events. The most interesting are the new bases introduced in the superstabile tetraloop of the hairpin ribozyme variants. Sequence alignment showed that PBD3 and 4 differ only by two bases and these are present in this tetraloop (positions 1 und 3). Since these two test systems also differ in their ability to form oligomers and RNA rings, it was concluded that these two positions are essential for programming ribozyme function towards circularization and/or oligomerisation

Oncoprotein 18 is necessary for malignant cell proliferation in bladder cancer cells and serves as a G3-specific non-invasive diagnostic marker candidate in urinary RNA.

BACKGROUND:Urine-based diagnostics indicated involvement of oncoprotein 18 (OP18) in bladder cancer. In cell culture models we investigated the role of OP18 for malignant cell growth. METHODS:We analyzed 113 urine samples and investigated two human BCa cell lines as a dual model: RT-4 and ECV-304, which represented differentiated (G1) and poorly differentiated (G3) BCa. We designed specific siRNA for down-regulation of OP18 in both cell lines. Phenotypes were characterized by cell viability, proliferation, and expression of apoptosis-related genes. Besides, sensitivity to cisplatin treatment was evaluated. RESULTS:Analysis of urine samples from patients with urothelial BCa revealed a significant correlation of the RNA-ratio OP18:uroplakin 1A with bladder cancer. High urinary ratios were mainly found in moderately to poorly differentiated tumors (grade G2-3) that were muscle invasive (stage T2-3), whereas samples from patients with more differentiated non-invasive BCa (G1) showed low OP18:UPK1A RNA ratios. Down-regulation of OP18 expression in ECV-304 shifted its phenotype towards G1 state. Further, OP18-directed siRNA induced apoptosis and increased chemo-sensitivity to cisplatin. CONCLUSIONS:This study provides conclusive experimental evidence for the link between OP18-derived RNA as a diagnostic marker for molecular staging of BCa in non-invasive urine-based diagnostics and the patho-mechanistic role of OP18 suggesting this gene as a therapeutic target

Technical research priorities for big data

To drive innovation and competitiveness, organisations need to foster the development and broad adoption of data technologies, value-adding use cases and sustainable business models. Enabling an effective data ecosystem requires overcoming several technical challenges associated with the cost and complexity of management, processing, analysis and utilisation of data. This chapter details a community-driven initiative to identify and characterise the key technical research priorities for research and development in data technologies. The chapter examines the systemic and structured methodology used to gather inputs from over 200 stakeholder organisations. The result of the process identified five key technical research priorities in the areas of data management, data processing, data analytics, data visualisation and user interactions, and data protection, together with 28 sub-level challenges. The process also highlighted the important role of data standardisation, data engineering and DevOps for Big Data

Genotype–Phenotype Relations for the Atypical Parkinsonism Genes: MDSGene Systematic Review

This Movement Disorder Society Genetic mutation database Systematic Review focuses on monogenic atypical parkinsonism with mutations in the ATP13A2, DCTN1, DNAJC6, FBXO7, SYNJ1, and VPS13C genes. We screened 673 citations and extracted genotypic and phenotypic data for 140 patients (73 families) from 77 publications. In an exploratory fashion, we applied an automated classification procedure via an ensemble of bootstrap-aggregated (“bagged”) decision trees to distinguish these 6 forms of monogenic atypical parkinsonism and found a high accuracy of 86.5% (95%CI, 86.3%–86.7%) based on the following 10 clinical variables: age at onset, spasticity and pyramidal signs, hypoventilation, decreased body weight, minimyoclonus, vertical gaze palsy, autonomic symptoms, other nonmotor symptoms, levodopa response quantification, and cognitive decline. Comparing monogenic atypical with monogenic typical parkinsonism using 2063 data sets from Movement Disorder Society Genetic mutation database on patients with SNCA, LRRK2, VPS35, Parkin, PINK1, and DJ-1 mutations, the age at onset was earlier in monogenic atypical parkinsonism (24 vs 40 years; P = 1.2647 × 10−12) and levodopa response less favorable than in patients with monogenic typical presentations (49% vs 93%). In addition, we compared monogenic to nonmonogenic atypical parkinsonism using data from 362 patients with progressive supranuclear gaze palsy, corticobasal degeneration, multiple system atrophy, or frontotemporal lobar degeneration. Although these conditions share many clinical features with the monogenic atypical forms, they can typically be distinguished based on their later median age at onset (64 years; IQR, 57–70 years). In conclusion, age at onset, presence of specific signs, and degree of levodopa response inform differential diagnostic considerations and genetic testing indications in atypical forms of parkinsonism

Franco-German position paper on "Speeding up industrial AI and trustworthiness"

The full benefit from using AI to generate value for businesses, societal wellbeing and the environment is still to be fully realised. To lower adoption barriers of Industrial AI, challenges on multiple levels (technical complexity, trustworthiness, industrialisation, data frameworks and infrastructures, etc.) need to be addressed to benefit from its full socio economic potential for economy, society and welfare. It is now time to foster the development of "industrial and trustworthy AI" and nurture European innovation and sovereignty ambitions and benefit society and leading European industries. In this position paper, a comprehensive industrial and trustworthy AI framework that clusters the priority area for AI research, innovation and deployment is introduced. It covers tools and methodologies that support the design, test, validation, verification, and maintainability of AI based functions and systems and addresses the development of AI based process and systems to demonstrate its integration into new products and services. Conformity assessment schemes, balancing innovation, business and European perspectives, are considered to connect risk management, functional andtrustworthiness requirements to industrial processes. In addition adequate standards supporting industrial AI and trustworthiness will play a central role. Implementing the industrial and trustworthy AI framework will require resources beyond the means of any European private stakeholders. Therefore, strong support from ecosystems, governments, and Europe might not be an option but necessary

Franco-German position paper on "Speeding up industrial AI and trustworthiness"

The full benefit from using AI to generate value for businesses, societal wellbeing and the environment is still to be fully realised. To lower adoption barriers of Industrial AI, challenges on multiple levels (technical complexity, trustworthiness, industrialisation, data frameworks and infrastructures, etc.) need to be addressed to benefit from its full socio economic potential for economy, society and welfare. It is now time to foster the development of "industrial and trustworthy AI" and nurture European innovation and sovereignty ambitions and benefit society and leading European industries. In this position paper, a comprehensive industrial and trustworthy AI framework that clusters the priority area for AI research, innovation and deployment is introduced. It covers tools and methodologies that support the design, test, validation, verification, and maintainability of AI based functions and systems and addresses the development of AI based process and systems to demonstrate its integration into new products and services. Conformity assessment schemes, balancing innovation, business and European perspectives, are considered to connect risk management, functional andtrustworthiness requirements to industrial processes. In addition adequate standards supporting industrial AI and trustworthiness will play a central role. Implementing the industrial and trustworthy AI framework will require resources beyond the means of any European private stakeholders. Therefore, strong support from ecosystems, governments, and Europe might not be an option but necessary