SUMMARY Enhancers control the correct temporal and cell type-specific activation of gene expression in higher eukaryotes. Knowing their properties, regulatory activity and targets is crucial to understand the regulation of differentiation and homeostasis. We use the FANTOM5 panel of samples covering the majority of human tissues and cell types to produce an atlas of active, in vivo transcribed enhancers. We show that enhancers share properties with CpG-poor mRNA promoters but produce bidirectional, exosome-sensitive, relatively short unspliced RNAs, the generation of which is strongly related to enhancer activity. The atlas is used to compare regulatory programs between different cells at unprecedented depth, identify disease-associated regulatory single nucleotide polymorphisms, and classify cell type-specific and ubiquitous enhancers. We further explore the utility of enhancer redundancy, which explains gene expression strength rather than expression patterns. The online FANTOM5 enhancer atlas represents a unique resource for studies on cell type-specific enhancers and gene regulation.

Regulated transcription controls the diversity, developmental pathways and spatial organization of the hundreds of cell types that make up a mammal. Using single-molecule cDNA sequencing, we mapped transcription start sites (TSSs) and their usage in human and mouse primary cells, cell lines and tissues to produce a comprehensive overview of mammalian gene expression across the human body. We find that few genes are truly ‘housekeeping’, whereas many mammalian promoters are composite entities composed of several closely separated TSSs, with independent cell-type-specific expression profiles. TSSs specific to different cell types evolve at different rates, whereas promoters of broadly expressed genes are the most conserved. Promoter-based expression analysis reveals key transcription factors defining cell states and links them to binding-site motifs. The functions of identified novel transcripts can be predicted by coexpression and sample ontology enrichment analyses. The functional annotation of the mammalian genome 5 (FANTOM5) project provides comprehensive expression profiles and functional annotation of mammalian cell-type-specific transcriptomes with wide applications in biomedical research.

A number of lanthanum strontium manganate/yttria-stabilized zirconia ͑LSM/YSZ͒ composite electrodes are produced with varying composition and processing parameters. The composites are investigated using impedance spectroscopy. General trends related to the oxygen reduction process are extracted from the impedance data. Literature concerning kinetic studies of LSM/YSZ electrodes and related systems is reviewed and compared to new experimental data. From this it is found that at least five processes affect the impedance. Going from high to low frequency, these processes are ͑i͒, ͑ii͒ two geometry-related contributions interpreted as transport across LSM/YSZ interfaces and through the YSZ of the composite, ͑iii͒ a process reflecting competitive reaction steps such as bond breaking and surface diffusion, (iv) gas diffusion in a stagnant gas layer above the electrode structure, and (v) an activation process ͑inductive͒ presumably located at the triple phase boundary of electrode, electrolyte, and gas phase.Lanthanum strontium manganate ͑LSM͒ is the classical cathode material for solid oxide fuel cells ͑SOFCs͒. 1 Addition of the electrolyte material yttria-stabilized zirconia ͑YSZ͒ to the cathode may improve the performance considerably compared to a simple porous LSM electrode. 2-3 To optimize the performance of such a composite electrode, it is important to gain knowledge about the oxygen reduction mechanism. Electrochemical impedance spectroscopy is used for this purpose, as it seems to be an efficient tool. The impedance of a composite cathode is complex. In a Nyquist plot, the impedance is composed of a number of more or less overlapping arcs each reflecting a physical or chemical process. Usually, it cannot be determined from a single impedance spectrum, how many physically relevant processes the impedance includes. Further, the impedance is affected by many features, which are not all known. These features are not all directly related to the oxygen reduction process. Therefore, it is necessary to perform many experiments varying parameters like temperature, oxygen partial pressure, overpotential and electrode geometry in order to interpret the impedance data using a simple physical or chemical model.The present paper attempts to summarize the knowledge about the O 2 /LSM/YSZ composite electrode gained through a large number of experiments performed on this system. Examples showing how the impedance is affected by processing and testing are given. Literature concerning kinetic studies of related electrode systems is reviewed. Some general trends related to the oxygen reduction mechanism are extracted from the presented impedance data and discussed in relation to literature. ExperimentalSample preparation.-Electrolyte substrates are produced from YSZ with 8 mol % Y 2 O 3 ͑TZ8Y, Tosoh Corporation, Japan͒. Threeelectrode electrochemical test cells are made using pellets, which are pressed and machined to the shape shown in Fig. 1a before sintering at 1600°C. The area of the working electrode is approximately 0.4 cm ...

Ntini et al. 3The non-protein-coding part of the human genome is pervasively transcribed into a large diversity of non-coding (nc) RNA 1 . A substantial fraction of this material derives from, or near, active gene promoters, that are producing a range of small-( 1-6 ) and long non-coding RNA (lncRNA) 7 . Indeed, it has been estimated that >60% of lncRNAs in human embryonic stem cells derive from promoters of active proteincoding genes 8 . Although some lncRNAs have reported functions, these species are generally kept at low abundance by cellular degradation activities 9,10 . For example, we previously coupled depletion of the major nuclear 3'-5' exonucleolytic activity, the RNA exosome, with tiling microarrays to reveal PROMPTs closely upstream of active human gene promoters 9 . PROMPTs are 5'capped, >100nt long and 3'end adenylated in the absence of exosome activity 11 . The mechanism underlying the efficient exosome-mediated suppression of these lncRNAs, while preserving the promoter-downstream mRNA, remains enigmatic.Here, we couple exosome-depletion to high-throughput 5'end-, 3'end-and regular RNA-sequencing (RNAseq) to create a genome-wide map of PROMPTs. Our results demonstrate that PROMPT transcription initiates antisense with respect to the downstream gene. We suggest that such initiating RNAPII, if stalled at a PROMPT-TSS proximal position, can elicit the production of previously reported TSSa-RNA.Sequence motifs around PROMPT 3'ends adhere to a pA site consensus and are significantly more abundant upstream than downstream of gene promoters. This provides a directional RNA output from human promoters by rapidly terminating antisense transcription and enforcing degradation of its RNA product. RESULTS PROMPTs initiate from bi-directional promoter activityTo obtain strand-specific and positional information of PROMPTs, we first subjected total RNA from HeLa cells, that had been treated with either a control (ctrl) eGFP siRNA or RRP40 siRNA ( Supplementary Fig. 1a), to regular RNA sequencing (RNAseq) as well as cap-selected RNA 5'end sequencing (Cap Analysis of Gene Expression (CAGE)). We focused our analysis on protein-coding genes and therefore considered reads mapping to the -3kb to +1kb regions of 2428 UCSC gene promoters, which were selected not to overlap any other annotated mRNAs. When aligned to the TSSs of these promoters, both RNAseq-and CAGE-data disclosed a strong presence of exosome-sensitive transcripts originating closely upstream of the gene TSS Ntini et al. 4(average peak CAGE position at -110bp) and commencing in the antisense direction relative to the neighboring mRNA TSS (Fig. 1a bottom panel, compare 'ctrl' and 'RRP40' plots). Only minor signal was detected in the sense direction of the same region. Whereas the abundance of antisense PROMPT (asPROMPT) CAGE tags increased by an average of 8-fold upon RRP40 depletion, the corresponding sense CAGE signals of the same region were largely unaffected (Fig. 1b, P<2e-16, twosided t-test). This predominant occurrence of asPROMPTs was also visi...

During the course of a lifetime normal human cells accumulate mutations. Here, using multiple samples from the same individuals we compared the mutational landscape in 29 anatomical structures from soma and the germline. Two ubiquitous mutational signatures, SBS1 and SBS5/40, accounted for the majority of acquired mutations in most cell types but their absolute and relative contributions varied substantially. SBS18, potentially reflecting oxidative damage, and several additional signatures attributed to exogenous and endogenous exposures contributed mutations to subsets of cell types. The .