Transcriptome analysis reveals a major impact of JAK protein tyrosine kinase 2 (Tyk2) on the expression of interferon-responsive and metabolic genes

<p>Abstract</p> <p>Background</p> <p>Tyrosine kinase 2 (Tyk2), a central component of Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling, has major effects on innate immunity and inflammation. Mice lacking Tyk2 are resistant to endotoxin shock induced by lipopolysaccharide (LPS), and Tyk2 deficient macrophages fail to efficiently induce interferon α/β after LPS treatment. However, how Tyk2 globally regulates transcription of downstream target genes remains unknown. Here we examine the regulatory role of Tyk2 in basal and inflammatory transcription by comparing gene expression profiles of peritoneal macrophages from Tyk2 mutant and wildtype control mice that were either kept untreated or exposed to LPS for six hours.</p> <p>Results</p> <p>Untreated Tyk2-deficient macrophages exhibited reduced expression of immune response genes relative to wildtype, in particular those that contain interferon response elements (IRF/ISRE), whereas metabolic genes showed higher expression. Upon LPS challenge, IFN-inducible genes (including those with an IRF/ISRE transcription factor binding-site) were strongly upregulated in both Tyk2 mutant and wildtype cells and reached similar expression levels. In contrast, metabolic gene expression was strongly decreased in wildtype cells upon LPS treatment, while in Tyk2 mutant cells the expression of these genes remained relatively unchanged, which exaggerated differences already present at the basal level. We also identified several 5'UR transcription factor binding-sites and 3'UTR regulatory elements that were differentially induced between Tyk2 deficient and wildtype macrophages and that have not previously been implicated in immunity.</p> <p>Conclusions</p> <p>Although Tyk2 is essential for the full LPS response, its function is mainly required for baseline expression but not LPS-induced upregulation of IFN-inducible genes. Moreover, Tyk2 function is critical for the downregulation of metabolic genes upon immune challenge, in particular genes involved in lipid metabolism. Together, our findings suggest an important regulatory role for Tyk2 in modulating the relationship between immunity and metabolism.</p

Solving a 50 year mystery of a missing OPA1 mutation: more insights from the first family diagnosed with autosomal dominant optic atrophy

Background: Up to the 1950s, there was an ongoing debate about the diversity of hereditary optic neuropathies, in particular as to whether all inherited optic atrophies can be ascribed to Leber's hereditary optic neuropathy (LHON) or represent different disease entities. In 1954 W. Jaeger published a detailed clinical and genealogical investigation of a large family with explicit autosomal dominant segregation of optic atrophy thus proving the existence of a discrete disease different from LHON, which is nowadays known as autosomal dominant optic atrophy (ADOA). Since the year 2000 ADOA is associated with genomic mutations in the OPA1 gene, which codes for a protein that is imported into mitochondria where it is required for mitochondrial fusion. Interestingly enough, the underlying mutation in this family has not been identified since then. Results: We have reinvestigated this family with the aim to identify the mutation and to further clarify the underlying pathomechanism. Patients showed a classical non-syndromic ADOA. The long term deterioration in vision in the two teenagers examined 50 years later is of particular note 5/20 to 6/120. Multiplex ligation probe amplification revealed a duplication of the OPA1 exons 7-9 which was confirmed by long distance PCR and cDNA analysis, resulting in an in-frame duplication of 102 amino acids. Segregation was verified in 53 available members of the updated pedigree and a penetrance of 88% was calculated. Fibroblast cultures from skin biopsies were established to assess the mitochondrial network integrity and to qualitatively and quantitatively study the consequences of the mutation on transcript and protein level. Fibroblast cultures demonstrated a fragmented mitochondrial network. Processing of the OPA1 protein was altered. There was no correlation of the OPA1 transcript levels and the OPA1 protein levels in the fibroblasts. Intriguingly an overall decrease of mitochondrial proteins was observed in patients' fibroblasts, while the OPA1 transcript levels were elevated. Conclusions: The thorough study of this family provides a detailed clinical picture accompanied by a molecular investigation of patients' fibroblasts. Our data show a classic OPA1-associated non-syndromic ADOA segregating in this family. Cell biological findings suggest that OPA1 is regulated by post-translational mechanisms and we would like to hypothesize that loss of OPA1 function might lead to impaired mitochondrial quality control. With the clinical, genetic and cell biological characterisation of a family described already more than 50 years ago, we span more than half a century of research in optic neuropathies

Transcriptome analysis reveals a major impact of JAK protein tyrosine kinase 2 (Tyk2) on the expression of interferon-responsive and metabolic genes

Background: Tyrosine kinase 2 (Tyk2), a central component of Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling, has major effects on innate immunity and inflammation. Mice lacking Tyk2 are resistant to endotoxin shock induced by lipopolysaccharide (LPS), and Tyk2 deficient macrophages fail to efficiently induce interferon alpha/beta after LPS treatment. However, how Tyk2 globally regulates transcription of downstream target genes remains unknown. Here we examine the regulatory role of Tyk2 in basal and inflammatory transcription by comparing gene expression profiles of peritoneal macrophages from Tyk2 mutant and wildtype control mice that were either kept untreated or exposed to LPS for six hours. Results: Untreated Tyk2-deficient macrophages exhibited reduced expression of immune response genes relative to wildtype, in particular those that contain interferon response elements (IRF/ISRE), whereas metabolic genes showed higher expression. Upon LPS challenge, IFN-inducible genes (including those with an IRF/ISRE transcription factor binding-site) were strongly upregulated in both Tyk2 mutant and wildtype cells and reached similar expression levels. In contrast, metabolic gene expression was strongly decreased in wildtype cells upon LPS treatment, while in Tyk2 mutant cells the expression of these genes remained relatively unchanged, which exaggerated differences already present at the basal level. We also identified several 5'UR transcription factor binding-sites and 3'UTR regulatory elements that were differentially induced between Tyk2 deficient and wildtype macrophages and that have not previously been implicated in immunity. Conclusions: Although Tyk2 is essential for the full LPS response, its function is mainly required for baseline expression but not LPS-induced upregulation of IFN-inducible genes. Moreover, Tyk2 function is critical for the downregulation of metabolic genes upon immune challenge, in particular genes involved in lipid metabolism. Together, our findings suggest an important regulatory role for Tyk2 in modulating the relationship between immunity and metabolism

EMI Security Architecture

This document describes the various architectures of the three middlewares that comprise the EMI software stack. It also outlines the common efforts in the security area that allow interoperability between these middlewares. The assessment of the EMI Security presented in this document was performed internally by members of the Security Area of the EMI project

Minimal information for studies of extracellular vesicles 2018 (MISEV2018):a position statement of the International Society for Extracellular Vesicles and update of the MISEV2014 guidelines

The last decade has seen a sharp increase in the number of scientific publications describing physiological and pathological functions of extracellular vesicles (EVs), a collective term covering various subtypes of cell-released, membranous structures, called exosomes, microvesicles, microparticles, ectosomes, oncosomes, apoptotic bodies, and many other names. However, specific issues arise when working with these entities, whose size and amount often make them difficult to obtain as relatively pure preparations, and to characterize properly. The International Society for Extracellular Vesicles (ISEV) proposed Minimal Information for Studies of Extracellular Vesicles (“MISEV”) guidelines for the field in 2014. We now update these “MISEV2014” guidelines based on evolution of the collective knowledge in the last four years. An important point to consider is that ascribing a specific function to EVs in general, or to subtypes of EVs, requires reporting of specific information beyond mere description of function in a crude, potentially contaminated, and heterogeneous preparation. For example, claims that exosomes are endowed with exquisite and specific activities remain difficult to support experimentally, given our still limited knowledge of their specific molecular machineries of biogenesis and release, as compared with other biophysically similar EVs. The MISEV2018 guidelines include tables and outlines of suggested protocols and steps to follow to document specific EV-associated functional activities. Finally, a checklist is provided with summaries of key points

Application of Two Novel Partial Least Squares-Based Regression Methods to the Analysis of Spectral Datasets

This work introduces two novel algorithms for multivariate regression: a partial least squares (PLS) variable selection method based on resampling and a PLS method using data transformation of the PLS weights (twPLS). The algorithms are tested on three spectral datasets (near-infrared and Raman) by predicting univariate response variables. The results are compared with the predictions of three other established methods comprising standard PLS, variable selection by sparse PLS (SPLS) and variable selection by variable importance in projection (VIP).Compared with the standard PLS method, the novel algorithms clearly improve predictions for one dataset and show slightly more accurate predictions for two other datasets.The two novel algorithms show comparable results with the SPLS and variable selection by VIP for all three datasets. These results demonstrate the effectiveness of the new approaches.</p

Application of Two Novel Partial Least Squares-Based Regression Methods to the Analysis of Spectral Datasets

This work introduces two novel algorithms for multivariate regression: a partial least squares (PLS) variable selection method based on resampling and a PLS method using data transformation of the PLS weights (twPLS). The algorithms are tested on three spectral datasets (near-infrared and Raman) by predicting univariate response variables. The results are compared with the predictions of three other established methods comprising standard PLS, variable selection by sparse PLS (SPLS) and variable selection by variable importance in projection (VIP).Compared with the standard PLS method, the novel algorithms clearly improve predictions for one dataset and show slightly more accurate predictions for two other datasets.The two novel algorithms show comparable results with the SPLS and variable selection by VIP for all three datasets. These results demonstrate the effectiveness of the new approaches

Reflections on Civil Liability Incentives in the Volkswagen Diesel Cases

Die Installation von Manipulationssoftware in Diesel-Pkw von Volkswagen führt zu unterschiedlichen Konsequenzen in den USA und in Deutschland. Die deutschen Käufer werden weniger oder gar nicht entschädigt, weil weder die staatliche Regulierung noch die privatrechtliche Haftung wirksam gegriffen hat. Jetzt besteht die Gefahr der Verjährung ihrer Ansprüche zum Jahresende. Die von der Großen Koalition in Aussicht gestellte Musterfeststellungsklage könnte jedoch noch rechtzeitig Abhilfe schaffen.The Volkswagen diesel emissions scandal is a case of market failure, because the harm from vehicle emissions to public health and to the environment was not internalised. This market failure has not been corrected by public regulation, nor by civil liability. There is also a regulatory failure, because the oversight of NOx emissions by the German automobile regulatory agency (Kraftfahrt-Bundesamt) was overly lenient. There is also a corporate governance failure within Volkswagen which made the fraud of installing manipulative software possible. And fi nally, there is dysfunctional law in Germany, which does not allow for class actions or similar types of lawsuits. The new government has announced the introduction of a new procedure (Musterfeststellungsklage) which would avoid this outcome