The serine/threonine kinase Prp4 of Schizosaccharomyces pombe improves the recognition of introns with weak splice sites

Die Genexpression in eukaryotischen Zellen gliedert sich in Transkription und verschiedene Prozessierungsschritte, wozu auch das prä-mRNA Spleißen gehört. Dabei werden aus der prä-mRNA die Introns herausgespleißt und die Exons miteinander verknüpft, so dass die reife mRNA entsteht. Das Herausspleißen der Introns wird vom Spleißosom durchgeführt, welches anhand der Exon1/5´-Spleißstelle, der Verzweigungssequenz und der 3´-Spleißstelle den Anfang und das Ende des Introns erkennt. In der vorliegenden Arbeit wurde die Funktion der Serin/Threonin Kinase Prp4 untersucht, die am Spleißprozess beteiligt ist. Dazu wurde eine analog-sensitive Kinase konstruiert, die sich durch Zugabe des ATP-analogen Inhibitors 1NM-PP1 hemmen lässt. Sowohl die Analyse einzelner Introns durch RT-PCR als auch eine Sequenzierung der Gesamt-RNA zeigen nach Inhibition der Prp4 Kinase, dass zwei Klassen von Introns existieren. Die eine Klasse der Introns wird nach Inhibition der Kinase weiter effizient gespleißt, während die andere Klasse nach Inhibition prä-mRNA akkumuliert. Die Introns, die die Aktivität der Prp4 Kinase benötigen um gespleißt zu werden, werden als Prp4-abhängige Introns bezeichnet. Und die Introns, die nach Inhibition weiter effizient gespleißt werden, werden Prp4-unabhängig genannt. Der Vergleich beider Intronklassen zeigt ausschließlich in den Sequenzen der Spleißstellen geringfügige Unterschiede. Um zu untersuchen, ob die Sequenz der Spleißstellen bezüglich der Prp4-Abhängigkeit eine Rolle spielt, wurden zwei Reportergene konstruiert. Dazu wurden die Gene res1, dessen Intron Prp4 abhängig gespleißt wird, und ppk8, welches unabhängig von der Kinase gespleißt wird, ausgewählt und in den leu1-Lokus von Schizosaccharomyces pombe integriert. Die Mutationsanalyse von Exon1/5´-SS und Verzweigungssequenz zeigt, dass durch Erhöhung der Komplementarität zur U1 und U2 snRNA des Spleißosoms ein Prp4-abhängiges Intron in ein Prp4-unabhängiges Intron umgewandelt werden kann. Wird die Komplementarität der snRNAs zur prä-mRNA verringert, kann aus einem Prp4-unabhängig gespleißten Intron ein Prp4 abhängiges werden. Das zeigt, dass die Aktivität der Prp4 Kinase notwendig ist, um Introns zu spleißen, deren Spleißstellen von der Konsensussequenz abweichen und damit als schwach bezeichnet werden.The gene expression of eukaryotic cells is divided into transcription and several processing steps. One of these processing steps is pre-mRNA splicing. In eukaryotic cells pre-mRNA consists of exons and introns and the introns have to be removed during the splicing reaction. The exons are joined together and the mature mRNA is formed. The splicing of introns is performed by the spliceosome which recognizes the beginning and the end of an intron via their exon1/5´-splice site, branch sequence and 3´ splice site. In this study the function of the serinethreonine kinase Prp4 which is involved in the splicing process is investigated. Therefore, an analog-sensitive kinase was constructed which can be inhibited using the ATP-analogue inhibitor 1NM-PP1. After Inhibition of the kinase several intron-containing genes were analyzed by RT-PCR. Moreover, the total RNA was sequenced. Both approaches revealed two classes of introns. One of these intron classes is still spliced after inhibition of the kinase while the other class accumulates pre-mRNA. The introns which need the activity of the kinase to be spliced out are called Prp4-dependent introns. And those that are spliced efficiently after inhibition of the kinase are named Prp4-independent introns. If both intron classes are compared only small differences between the sequences of the splice sites can be observed. Two reporter genes were constructed to analyse if the sequence of the splice sites play a role in Prp4-dependency. Therefore, the gene res1 which contains a Prp4-dependent intron and the gene ppk8 which contains a Prp4-independent intron were integrated into the leu1-locus of Schizosaccharomyces pombe. The analysis of different mutations in exon1/5´-SS and branch sequence showed that increasing the complementarity to the snRNAs U1 and U2 of the spliceosome can change a Prp4-dependent intron in an independent one. Decreasing the complementarity of a Prp4-independent intron leads to Prp4 dependency. That shows that the activity of the Prp4 kinase is necessary for splicing of introns whose splice sites differ from the consensus sequence. Those are called weak splice sites

Circular economy for durable products and materials : the recycling of plastic building products in Germany ; status quo, potentials and recommendations

The construction sector is the second largest area for the application for plastics. Due to the long life times of construction products, the implementation of the circular economy faces its own challenges. To investigate this challenge, the study covers a market study for Germany, voluntary take-back and recycling schemes of construction products, as well as the use of plastic recyclates in construction products. In addition, plastic packaging of construction products is covered. Opportunities and barriers to the use of recycled plastics in construction products are derived from the intersection of available technologies, recyclate supply, and technical requirements for construction products. The report concludes with recommendations to various stakeholders on how to promote the use of recyclates in construction products and their packaging. Important points here are the introduction of a recyclate quota for films as construction product packaging and the description of recycling possibilities and recyclate content in the technical documentation of construction products

Clockwise rotation of the Baoshan block due to SSE-ward extrusion of Tibetan crust

Abstract HKT-ISTP 2013 A

Promoting the high-quality recycling of plastics from demolition waste and enhancing the use of recycled materials in construction products in accordance with the European plastics strategy : final report

This study presents in detail: the use of plastic products and the opportunities for recyclate use in the construction sector, quantities of plastic used, take-back systems, recycling techniques, current recyclate use and plastic construction product packaging.Potentials for increasing high-quality recyclate use were identified. Existing hurdles and options for action for industry and politics are presented. Current recyclate use as well as its potential use are strongly dependent on the application area of plastics. The biggest hurdles for the use of recycled materials are product life time, dismantling and technical requirements

ENTREPRENEURSHIP AND INNOVATION HUB AT A PRIVATE UNIVERSITY IN RIO GRANDE DO SUL

The proposed hub of Academic Activities in the areas of Innovation and Entrepreneurship for teaching undergraduate students originated in the strategic project of the University entitled "Tecnosinos Project - São Leopoldo Tech Park". The main objective of this project is to make Unisinos University a technology-fostering agent, conducting applied research in Entrepreneurship and Innovation. This paper will discuss the importance of the entrepreneurship education and the successful partnership between Unisinos and Tecnosinos Tech Park on an innovative program that has positively influenced a large number of students since 2011, fostering the birth of new startups. The methodology of this article is related to a case study originated from a qualitative applied research with secondary data collection from register and from academic curricula. Additionally this work took primary data collection through interview with professors and with Unitec manager, who is responsible of this project from Tecnosinos side. Results from this study shows increasing number of students participants of different kind of activities. A part from education, some activities focused on entrepreneurship promotion, such as Warm up, Roser Award and Innomarathon, that resulted in more than six projects that became startups at Unitec Incubator. Keywords: Entrepreneurship. Innovation. Hub. Connection. Education.   RESUMO A proposta de criação do eixo de atividades acadêmicas nas áreas de Inovação e Empreendedorismo, para ensinar esses temas aos alunos da Graduação, originou-se no projeto estratégico da Universidade denominado “Projeto Tecnosinos – Parque Tecnológico São Leopoldo”. O principal objetivo desse projeto é promover a Unisinos como um agente de fomento ao desenvolvimento tecnológico, conduzindo pesquisa aplicada em Inovação e Empreendedorismo. Este artigo discute a importância da educação empreendedora e a parceria de sucesso entre a Unisinos e o Tecnosinos – Parque Tecnológico de São Leopoldo, como um programa inovador que influencia positivamente um grande número de estudantes, desde 2011, promovendo o surgimento de novas startups. Este artigo é um estudo de caso originado de uma pesquisa qualitativa baseada em dados secundários obtidos dos registros do Tecnosinos e de análise das ementas das disciplinas de empreendedorismo. Adicionalmente foram feitas algumas entrevistas com professores do eixo e com o gerente da Incubadora Unitec, responsável por esse projeto da parte do Tecnosinos. Os resultados desse estudo mostram que há um número crescente de estudantes participantes em diferentes atividades relacionados ao eixo. Além da educação empreendedora, algumas atividades estão focadas em promover o empreendedorismo, tais como Warm up, o Prêmio Roser e o Innomarathon que resultaram em mais de seis projetos que se tornaram startups na Incubadora Unitec. Palavras-chave: Empreendedorismo. Inovação. Eixo. Conexões. Educação

Rationale for Combining Radiotherapy and Immune Checkpoint Inhibition for Patients With Hypoxic Tumors

In order to compensate for the increased oxygen consumption in growing tumors, tumors need angiogenesis and vasculogenesis to increase the supply. Insufficiency in this process or in the microcirculation leads to hypoxic tumor areas with a significantly reduced pO2, which in turn leads to alterations in the biology of cancer cells as well as in the tumor microenvironment. Cancer cells develop more aggressive phenotypes, stem cell features and are more prone to metastasis formation and migration. In addition, intratumoral hypoxia confers therapy resistance, specifically radioresistance. Reactive oxygen species are crucial in fixing DNA breaks after ionizing radiation. Thus, hypoxic tumor cells show a two- to threefold increase in radioresistance. The microenvironment is enriched with chemokines (e.g., SDF-1) and growth factors (e.g., TGFβ) additionally reducing radiosensitivity. During recent years hypoxia has also been identified as a major factor for immune suppression in the tumor microenvironment. Hypoxic tumors show increased numbers of myeloid derived suppressor cells (MDSCs) as well as regulatory T cells (Tregs) and decreased infiltration and activation of cytotoxic T cells. The combination of radiotherapy with immune checkpoint inhibition is on the rise in the treatment of metastatic cancer patients, but is also tested in multiple curative treatment settings. There is a strong rationale for synergistic effects, such as increased T cell infiltration in irradiated tumors and mitigation of radiation-induced immunosuppressive mechanisms such as PD-L1 upregulation by immune checkpoint inhibition. Given the worse prognosis of patients with hypoxic tumors due to local therapy resistance but also increased rate of distant metastases and the strong immune suppression induced by hypoxia, we hypothesize that the subgroup of patients with hypoxic tumors might be of special interest for combining immune checkpoint inhibition with radiotherapy

Förderung einer hochwertigen Verwertung von Kunststoffen aus Abbruchabfällen sowie der Stärkung des Rezyklateinsatzes in Bauprodukten im Sinne der europäischen Kunststoffstrategie

In dieser Studie werden detailliert dargestellt: die Verwendung von Kunststoffprodukten und Möglichkeiten des Rezyklateinsatzes im Baubereich, verwendete Kunststoffmengen, Rücknahmesysteme, Verwertungstechniken, aktueller Rezyklateneinsatz und Bauproduktverpackungen aus Kunststoff. Potentiale zur Steigerung des hochwertigen Rezyklateinsatzes wurden identifiziert und bestehende Hürden und Handlungsoptionen für Politik und Wirtschaft dargestellt. Aktueller Rezyklateinsatz sowie technische und mengenmäßige Potentiale sind stark abhängig vom Einsatzbereich der Kunststoffe. Die größten Hürden für einen Rezyklateinsatz sind dabei Produktlebensdauer, Rückbaufähigkeit und technische Anforderungen

Plastin 3 is upregulated in iPSC-derived motoneurons from asymptomatic SMN1-deleted individuals

Spinal muscular atrophy (SMA) is a devastating motoneuron (MN) disorder caused by homozygous loss of SMN1. Rarely, SMN1-deleted individuals are fully asymptomatic despite carrying identical SMN2 copies as their SMA III-affected siblings suggesting protection by genetic modifiers other than SMN2. High plastin 3 (PLS3) expression has previously been found in lymphoblastoid cells but not in fibroblasts of asymptomatic compared to symptomatic siblings. To find out whether PLS3 is also upregulated in MNs of asymptomatic individuals and thus a convincing SMA protective modifier, we generated induced pluripotent stem cells (iPSCs) from fibroblasts of three asymptomatic and three SMA III-affected siblings from two families and compared these to iPSCs from a SMA I patient and control individuals. MNs were differentiated from iPSC-derived small molecule neural precursor cells (smNPCs). All four genotype classes showed similar capacity to differentiate into MNs at day 8. However, SMA I-derived MN survival was significantly decreased while SMA III- and asymptomatic-derived MN survival was moderately reduced compared to controls at day 27. SMN expression levels and concomitant gem numbers broadly matched SMN2 copy number distribution; SMA I presented the lowest levels, whereas SMA III and asymptomatic showed similar levels. In contrast, PLS3 was significantly upregulated in mixed MN cultures from asymptomatic individuals pinpointing a tissue-specific regulation. Evidence for strong PLS3 accumulation in shaft and rim of growth cones in MN cultures from asymptomatic individuals implies an important role in neuromuscular synapse formation and maintenance. These findings provide strong evidence that PLS3 is a genuine SMA protective modifier