Janus—a comprehensive tool investigating the two faces of transcription

Motivation: Protocols to generate strand-specific transcriptomes with next-generation sequencing platforms have been used by the scientific community roughly since 2008. Strand-specific reads allow for detection of antisense events and a higher resolution of expression profiles enabling extension of current transcript annotations. However, applications making use of this strandedness information are still scarce. Results: Here we present a tool (Janus), which focuses on the identification of transcriptional active regions in antisense orientation to known and novel transcribed elements of the genome. Janus can compare the antisense events of multiple samples and assigns scores to identify mutual expression of either transcript in a sense/antisense pair, which could hint to regulatory mechanisms. Janus is able to make use of single-nucleotide variant (SNV) and methylation data, if available, and reports the sense to antisense ratio of regions in the vicinity of the identified genetic and epigenetic variation. Janus interrogates positions of heterozygous SNVs to identify strand-specific allelic imbalance. Availability: Janus is written in C/C++ and freely available at http://www.ikmb.uni-kiel.de/janus/janus.html under terms of GNU General Public License, for both, Linux and Windows 64×. Although the binaries will work without additional downloads, the software depends on bamtools (https://github.com/pezmaster31/bamtools) for compilation. A detailed tutorial section is included in the first section of the supplemental material and included as brief readme.txt in the tutorial archive. Contact: [email protected] or [email protected] Supplementary information: Supplementary data are available at Bioinformatics onlin

JAK-STAT inhibition impairs K-RAS-driven lung adenocarcinoma progression

Oncogenic KRAS has been difficult to target and currently there is no KRASbased targeted therapy available for patients suffering from KRASdriven lung adenocarcinoma (AC). Alternatively, targeting KRASdownstream effectors, KRAScooperating signaling pathways or cancer hallmarks, such as tumorpromoting inflammation, has been shown to be a promising therapeutic strategy. Since the JAKSTAT pathway is considered to be a central player in inflammationmediated tumorigenesis, we investigated here the implication of JAKSTAT signaling and the therapeutic potential of JAK1/2 inhibition in KRASdriven lung AC. Our data showed that JAK1 and JAK2 are activated in human lung AC and that increased activation of JAKSTAT signaling correlated with disease progression and KRAS activity in human lung AC. Accordingly, administration of the JAK1/2 selective tyrosine kinase inhibitor ruxolitinib reduced proliferation of tumor cells and effectively reduced tumor progression in immunodeficient and immunocompetent mouse models of KRASdriven lung AC. Notably, JAK1/2 inhibition led to the establishment of an antitumorigenic tumor microenvironment, characterized by decreased levels of tumorpromoting chemokines and cytokines and reduced numbers of infiltrating myeloid derived suppressor cells, thereby impairing tumor growth. Taken together, we identified JAK1/2 inhibition as promising therapy for KRASdriven lung AC.(VLID)510233

The functional characterisation of the C-type lectin : Clecsf8

Includes bibliographical references (leaves 147-157).Clecsf8 is a poorly characterised member of the "Dectin-2 cluster" and was originally thought to be expressed exclusively by macrophages. In this study it was demonstrated that Clecsf8 is primarily expressed by peripheral blood neutrophils and monocytes

Quantitative methods for profiling dynamic chromatin features

Living systems, from entire organisms down to the single cells constituting them are dynamic entities that continuously adapt and respond to their local environment. Cells achieve this through gene expression programs derived from static information encoded in the DNA made dynamic through chemical modifications at the chromatin level, collectively termed the epigenome. Numerous epigenetic regulators have been implicated in early embryonic developmental transitions and pluripotency. Ex vivo, the different states of pluripotency can be recapitulated by embryonic stem cells (ESCs) grown in defined media conditions. Many developmental gene promoters in ESCs display co-occurrence of the activating histone H3 lysine 4 trimethylation (H3K4me3) mark and the repressive H3K27me3 mark. This distinctive bivalent signature is considered to poise expression, allowing timely resolution to an active or inactive state depending on the signal. The distribution of histone modifications and chromatin-associated factors across the genome can be mapped using chromatin immunoprecipitation followed by next-generation sequencing (ChIP-seq). However, traditional ChIP-seq methods fail to quantitatively profile the nuanced global and local epigenetic rewiring that takes place in key developmental stages. This thesis addresses this limitation through the development of a quantitative multiplexed ChIP-seq technology: MINUTE (multiplexed indexed unique molecule T7 amplification end to end sequencing) ChIP. Across the three papers included in this thesis, we reveal the underpinnings of chromatin state dynamics in early mouse and human embryonic development by employing MINUTE ChIP. In Paper I, we first show that MINUTE ChIP enables accurate quantitative comparisons over a wide linear range. By employing it to characterize mouse ESCs grown in 2i and serum conditions, we find that the 2i naïve state is characterized by high global levels of H3K27me3 and low H3K4me3. At bivalent promoters, we observe that while H3K27me3 levels are stably maintained between serum and 2i, H3K4me3 levels are higher in the serum condition. Through quantitative epigenome profiling, in Paper II we find that naïve human ESCs also have broad global gain of Polycomb repressive complex 2 (PRC2)-mediated H3K27me3 and define a previously unrecognized, naïve-specific set of bivalent promoters. Bulk and single-cell transcriptomics confirmed that naïve bivalency maintains key trophectoderm and mesoderm transcription factors in a transcriptionally poised state which is resolved to an active state upon depletion of H3K27me3. Therefore, we discovered that PRC2-mediated repression provides a highly adaptive mechanism to restrict lineage potential during early human development. In paper III we show how quantitative RNA polymerase II occupancy profiles generated by MINUTE ChIP can be integrated with transient transcriptomics data to unravel genome wide transcriptional kinetics in three mESCs pluripotent states: naïve, ground and paused. Taken together, this thesis provides compelling evidence for a broad H3K27me3 hypermethylation of the genome in both naïve mouse and human ESCs and the basis for substantially revising the model for bivalency during embryonic developmen

Signal Balance as a Pluripotency Determinant: In vitro modeling of in vivo pluripotency states with WNT, FGF and BMP

This thesis enriches the body of knowledge around pluripotency by describing a novel pluripotent state. It deepens our understanding of cell biology, the processes establishing pluripotency and provides new insights into improving the process of artificial generation of pluripotent cells

Identifikation funktioneller genetischer Varianten in entzündlichen Darmerkrankungen durch Genom- und Transkriptomsequenzierung

Dissertation dealing with 1) the analysis of the whole genome and transcriptome of a family with a severe case with early onset of Crohn's Disease and 2) the development of an application for the genome-wide analysis of antisense transcription in Next-Generation-Sequencing.Dissertation, die sich mit der Analyse des gesamten Genoms und Transkriptoms einer Familie mit schwerem Fall und frühem Ausbruch von Morbus Crohn beschäftigt, so wie mit der Entwicklung einer Applikation zur genomweiten Analyse von Antisense-Transkription durch Next-Generation-Sequencing

The Struggle for Law: legal strategies, environmental struggles and climate actions in Italy

The legal field constitutes both an advantageous and restrictive tool for social struggles. Current movements trust law and litigation as beneficial instruments. Nevertheless, legal strategies are deployed against protestors too. This article stems from processes of criminalization and unsuccessful “struggle for law” towards cases where legal strategies are embraced by social movements to demonstrate the Janus-faced nature of the legal field. The focus is on Italy, and is based on two-year judicial ethnography on the No TAV movement and fieldwork as well as documental analysis on the anti-Trans-Adriatic Pipeline (TAP) mobilization. Hence, the analysis focuses on a “positive” case selection, namely the most relevant climate litigation strategies pursued worldwide, with a special focus on Giudizio Universale, the first Italian action. The goal is to grasp both the conflicting aspects and the resourceful nature of law through empirical cases and case law to evaluate potential successful practices for current social movements

Phenomenological Reflections of Trauma Survivors on Healing through Yoga

A hermeneutic phenomenological approach was used to explore the effects of trauma on occupational participation and well-being and how yoga impacts healing. Three trauma survivors with yoga practices consisting of asana (postures), pranayama (breath control), and meditation shared their stories. Interviews yielded compelling accounts of how yoga has helped participants manage the evolving effects of trauma and open towards embodied healing. The major theme embodiment was arrived at with constituent themes: choices, opening towards possibilities, connection, the power of breath, yoga as a microcosm. Findings suggest opportunity for integration of yoga and occupational therapy in a holistic approach to trauma