Project description:Asthma is a complex syndrome associated with episodic decompensations provoked by aeroaller-gen exposures. The underlying pathophysiological states driving exacerbations are latent in the resting state and do not adequately inform biomarker-driven therapy. A better understanding of the pathophysiological pathways driving allergic exacerbations is needed. We hypothesized that disease-associated pathways could be identified in humans by unbiased metabolomics of bron-choalveolar fluid (BALF) during the peak inflammatory response provoked by a bronchial aller-gen challenge. We analyzed BALF metabolites in samples from 12 volunteers who underwent segmental bronchial antigen provocation (SBP-Ag). Metabolites were quantified using liquid chromatography-tandem mass spectrometry (LC–MS/MS) followed by pathway analysis and cor-relation with airway inflammation. SBP-Ag induced statistically significant changes in 549 fea-tures that mapped to 72 uniquely identified metabolites. From these features, two distinct induci-ble metabolic phenotypes were identified by the principal component analysis, partitioning around medoids (PAM) and k-means clustering. Ten index metabolites were identified that in-formed the presence of asthma-relevant pathways, including unsaturated fatty acid produc-tion/metabolism, mitochondrial beta oxidation of unsaturated fatty acid, and bile acid metabolism. Pathways were validated using proteomics in eosinophils. A segmental bronchial allergen chal-lenge induces distinct metabolic responses in humans, providing insight into pathogenic and pro-tective endotypes in allergic asthma.
Project description:We used ATLAS-seq to comprehensively map the genomic location of LINE-1 elements belonging to the youngest and potentially polymorphic subfamily (L1HS-Ta). This was performed in a panel of 12 human primary or transformed cell lines (BJ, IMR90, MRC5, H1, K562, HCT116, HeLa S3, HepG2, MCF7, HEK-293, HEK-293T, 2102Ep). In brief, ATLAS-seq relies on the random mechanical fragmentation of the genomic DNA to ensure high-coverage, ligation of adapter sequences, suppression PCR-amplification of L1HS-Ta element junctions, and Ion Torrent sequencing using single-end 400 bp read chemistry. A notable aspect of ATLAS-seq is that we can obtain both L1 downstream and upstream junctions (3'- and 5'-ATLAS-seq libraries, respectively), for full-length L1 elements. Note that a 10-nt sample-specific barcode has been removed at the 5' end of the reads in the .fastq files upon demultiplexing. This was achieved using cutadapt v1.9.2.dev0 (with the following parameters: -e 0.1 -q 10 -m 25 -g <barcode_name>=^<barcode_sequence>)
Project description:MicroRNAs are important negative regulators of protein coding gene expression, and have been studied intensively over the last few years. To this purpose, different measurement platforms to determine their RNA abundance levels in biological samples have been developed. In this study, we have systematically compared 12 commercially available microRNA expression platforms by measuring an identical set of 20 standardized positive and negative control samples, including human universal reference RNA, human brain RNA and titrations thereof, human serum samples, and synthetic spikes from homologous microRNA family members. We developed novel quality metrics in order to objectively assess platform performance of very different technologies such as small RNA sequencing, RT-qPCR and (microarray) hybridization. We assessed reproducibility, sensitivity, quantitative performance, and specificity. The results indicate that each method has its strengths and weaknesses, which helps guiding informed selection of a quantitative microRNA gene expression platform in function of particular study goals.
Project description:This study consists of 10 whole genome RNA-seq profiles which have been generated from blood samples collected from ten different volunteers in the Personal Genome Project UK
Project description:C2C12 myoblast is a model that has been used extensively to study the process of skeletal muscle differentiation. Proteomics has advanced our understanding of skeletal muscle biology and the process of myogenesis. However, there is still no deep coverage of C2C12 myoblast proteome, which is important for the understanding of key drivers for the differentiation of skeletal muscle cells. Here, we conducted a multi-dimensional proteome profiling with TiSH strategy to get a comprehensive analysis of proteome, phosphoproteome and N-linked sialylated glycoproteome of C2C12 myoblasts. A total of 8313 protein groups were identified in C2C12 myoblasts, including 7827 protein groups from non-modified peptides, 3803 phosphoproteins and 977 formerly N-linked sialylated glycoproteins.
Project description:Genome-wide identification of transcription factor (TF) binding sites in the genome of the fission yeast Schizosaccharomyces pombe. The ChIP-nexus method was used. TFs included were: Cbf11-TAP and Cbf12-TAP (and their DBM mutants with impaired DNA binding), TAP-Mga2, and Fkh2-TAP (as an irrelevant control TF). IPs from an untagged WT strain were also analyzed. Cbf11-related IPs were performed from exponential cultures, while Cbf12-related IPs were performed from stationary cultures. YES complex medium was used for all cultivations.