SummaryAs the premier model organism in biomedical research, the laboratory mouse shares the majority of protein-coding genes with humans, yet the two mammals differ in significant ways. To gain greater insights into both shared and species-specific transcriptional and cellular regulatory programs in the mouse, the Mouse ENCODE Consortium has mapped transcription, DNase I hypersensitivity, transcription factor binding, chromatin modifications, and replication domains throughout the mouse genome in diverse cell and tissue types. By comparing with the human genome, we not only confirm substantial conservation in the newly annotated potential functional sequences, but also find a large degree of divergence of other sequences involved in transcriptional regulation, chromatin state and higher order chromatin organization. Our results illuminate the wide range of evolutionary forces acting on genes and their regulatory regions, and provide a general resource for research into mammalian biology and mechanisms of human diseases.

Summary The three-dimensional configuration of DNA is integral to all nuclear processes in eukaryotes, yet our knowledge of the chromosome architecture is still limited. Genome-wide chromosome conformation capture studies have uncovered features of chromatin organization in cultured cells, but genome architecture in human tissues has yet to be explored. Here, we report the most comprehensive survey to date of chromatin organization in human tissues. Through integrative analysis of chromatin contact maps in 21 primary human tissues and cell types, we found topologically associating domains highly conserved in different tissues. We also discover genomic regions that exhibit unusually high levels of local chromatin interactions. These frequently interacting regions (FIREs) are enriched for super-enhancers and are near tissue-specifically expressed genes. They display strong tissue-specificity in local chromatin interactions. Additionally, FIRE formation is partially dependent on CTCF and the Cohesin complex. We further show that FIREs can help annotate function of non-coding sequence variants.

SummaryUnderstanding the diversity of human tissues is fundamental to disease and requires linking genetic information, which is identical in most of an individual’s cells, with epigenetic mechanisms that could play tissue-specific roles. Surveys of DNA methylation in human tissues have established a complex landscape including both tissue-specific and invariant methylation patterns1,2. Here we report high coverage methylomes that catalogue cytosine methylation in all contexts for the major human organ systems, integrated with matched transcriptomes and genomic sequence. By combining these diverse data types with each individuals’ phased genome3, we identified widespread tissue-specific differential CG methylation (mCG), partially methylated domains, allele-specific methylation and transcription, and the unexpected presence of non-CG methylation (mCH) in almost all human tissues. mCH correlated with tissue-specific functions, and using this mark, we made novel predictions of genes that escape X-chromosome inactivation in specific tissues. Overall, DNA methylation in multiple genomic contexts varies substantially among human tissues.

In Drosophila, the neuropeptide pigment-dispersing factor (PDF) is required to maintain behavioral rhythms under constant conditions. To understand how PDF exerts its influence, we performed time-series immunostainings for the PERIOD protein in normal and pdf mutant flies over 9 d of constant conditions. Without pdf, pacemaker neurons that normally express PDF maintained two markers of rhythms: that of PERIOD nuclear translocation and its protein staining intensity. As a group, however, they displayed a gradual dispersion in their phasing of nuclear translocation. A separate group of non-PDF circadian pacemakers also maintained PERIOD nuclear translocation rhythms without pdf but exhibited altered phase and amplitude of PERIOD staining intensity. Therefore, pdf is not required to maintain circadian protein oscillations under constant conditions; however, it is required to coordinate the phase and amplitude of such rhythms among the diverse pacemakers. These observations begin to outline the hierarchy of circadian pacemaker circuitry in the Drosophila brain.Key words: pigment-dispersing factor; circadian rhythm; Drosophila; lateral neurons; nuclear accumulation; period IntroductionThe organizing principles for the neuronal networks underlying circadian oscillations are essentially unknown. Which cells are the critical oscillators for particular output functions, what is their hierarchical organization, and how are synchronizing signals coordinated among pacemaker groups to provide coherent circadian output? In the Drosophila brain, ϳ100 pacemaker neurons are defined by expression of period ( per) and other genes essential for circadian rhythmicity (Kaneko and Hall, 2000). These clock cells are divided into the lateral (LN) and dorsal (DN) neural groups (Helfrich-Förster, 2003). Mosaic analysis suggests that LNs, but not DNs, are necessary to establish locomotor rhythms (Frisch et al., 1994;Vosshall and Young, 1995). LNs are segregated into distinct dorsal (LN d ) and ventral (LN v ) groups; the latter is divided into small and large subgroups. The LN v clock neurons express the neuropeptide pigment-dispersing factor ( pdf ) (Helfrich-Förster, 1998). Genetic ablation of the entire LN v group produces a syndrome similar to that observed in pdf 01 mutant flies (Renn et al., 1999). Such flies anticipate light-to-dark transition events but are phase advanced; in constant darkness, ϳ70% lose their locomotor rhythms, and the remainder display only weak rhythms.The rhythmic nature of single pacemaker cells has traditionally been ascribed to individual cell properties (Michel et al., 1993;Welsh et al., 1995;Liu et al., 1997;Herzog et al., 1998). Recent evidence, however, suggests that interneuronal communication may be required to sustain basic molecular rhythms.Manipulations that alter pacemaker membrane excitability or disrupt transmitter signaling between pacemakers result in severe dampening of molecular rhythms (Harmar et al., 2002;Nitabach et al., 2002;Colwell et al., 2003;Lee et al., 2003). Likewise, Peng et al. (...

Although the similarities between humans and mice are typically highlighted, morphologically and genetically, there are many differences. To better understand these two species on a molecular level, we performed a comparison of the expression profiles of 15 tissues by deep RNA sequencing and examined the similarities and differences in the transcriptome for both protein-coding and -noncoding transcripts. Although commonalities are evident in the expression of tissue-specific genes between the two species, the expression for many sets of genes was found to be more similar in different tissues within the same species than between species. These findings were further corroborated by associated epigenetic histone mark analyses. We also find that many noncoding transcripts are expressed at a low level and are not detectable at appreciable levels across individuals. Moreover, the majority lack obvious sequence homologs between species, even when we restrict our attention to those which are most highly reproducible across biological replicates. Overall, our results indicate that there is considerable RNA expression diversity between humans and mice, well beyond what was described previously, likely reflecting the fundamental physiological differences between these two organisms.transcriptome | epigenome | species comparison | noncoding transcripts T he mouse has served as a valuable model organism for human biology and disease. It is widely assumed that biochemical, cellular, and developmental pathways in the mouse are highly conserved with humans and that many processes are clearly preserved at a molecular and genetic level. Moreover, recent detailed studies have examined gene expression in a limited number of tissues in humans and mice. These studies have indicated that gene expression is often conserved and is more similar between the comparable tissues of different organisms rather than within tissues of the same organism. In contrast, the transcript isoform repertoire was found to be markedly different between species (1, 2). Gene Expression Is More Similar Among Tissues Within a Species Than Between Corresponding Tissues of the Two SpeciesTo examine the similarities between humans and mice in much greater detail, we produced RNA-seq data from 13 human tissues [as part of the Encyclopedia Of DNA Elements (ENCODE)], another 11 human tissues [as part of the Roadmap Epigenomics Mapping Consortium (REMC) (3)], and 13 mouse tissues (for mouse ENCODE). We also included in our analysis other data from mouse ENCODE and the Illumina Human BodyMap 2.0 (HBM) (SI Materials and Methods). Sequencing was performed to a depth of 11,313,824-166,188,101 mappable reads (median of 68,399,538 with and an interquartile range of 31,557,836,199). In total, our analysis used 93 datasets encompassing the most tissue-diverse RNA-seq dataset to date spanning several major projects. Thirteen of the mouse and human orthologous datasets were produced by the same laboratory. For our analysis regarding noncoding transcripts, we incorporated an ad...

Summary To broaden our understanding of the evolution of gene regulation mechanisms, we generated occupancy profiles for 34 orthologous transcription factors (TFs) in human-mouse erythroid progenitor, lymphoblast, and embryonic stem cell lines. By combining the genome-wide TF occupancy repertoires, associated epigenetic signals, and TF co-association patterns, we deduced several evolutionary principles of gene regulatory features operating since the mouse and human lineages diverged. The genomic distribution profiles, primary binding motifs, chromatin states, and DNA methylation preferences are well conserved for TF occupied sequences (TF OSs). However, the extent to which orthologous DNA segments are bound by orthologous TFs varies both among TFs and with genomic location: binding at promoters is more highly conserved than binding at distal elements. Importantly, occupancy conserved TF OSs tend to be pleiotropic; they function in multiple tissues and also co-associate with multiple TFs. Single nucleotide variants (SNVs) at sites with potential regulatory functions are enriched in occupancy conserved TF OSs.

Allelic differences between the two homologous chromosomes can affect the propensity of inheritance in humans; however, the extent of such differences in the human genome has yet to be fully explored. Here, for the first time, we delineate allelic chromatin modifications and transcriptomes amongst a broad set of human tissues, enabled by a chromosome-spanning haplotype reconstruction strategy1. The resulting masses of haplotype-resolved epigenomic maps reveal extensive allelic biases in both chromatin state and transcription, which show considerable variation across tissues and between individuals, and allow us to investigate cis-regulatory relationships between genes and their control sequences. Analyses of histone modification maps also uncover intriguing characteristics of cis-regulatory elements and tissue-restricted activities of repetitive elements. The rich datasets described here will enhance our understanding of the mechanisms of how cis-regulatory elements control gene expression programs.

We measured daily gene expression in heads of control and period mutant Drosophila by using oligonucleotide microarrays. In control flies, 72 genes showed diurnal rhythms in light-dark cycles; 22 of these also oscillated in free-running conditions. The period gene significantly influenced the expression levels of over 600 nonoscillating transcripts. Expression levels of several hundred genes also differed significantly between control flies kept in light-dark versus constant darkness but differed minimally between per 01 flies kept in the same two conditions. Thus, the period-dependent circadian clock regulates only a limited set of rhythmically expressed transcripts. Unexpectedly, period regulates basal and light-regulated gene expression to a very broad extent.F orward genetic screens in Drosophila melanogaster have identified at least eight genes [period (per), timeless (tim), cycle (cyc), clock (Clk), vrille (vri), doubletime, cryptochrome (cry), and shaggy] necessary for the normal functioning of the circadian time-keeping system. Null mutations in most of these genes render flies behaviorally arrhythmic in constant conditions, but they otherwise have minimal morphologic phenotype (1). A model for the mechanism by which specific gene products give rise to a stable clock mechanism has been formulated over the past 10 years (2, 3). These clock genes appear to function in a time-delayed transcription-translation feedback loop. A rhythmically expressed subset of the core clock genes (per, tim, and Clk) and a nonrhythmically expressed core clock gene (cyc) are thought to function as the state variables of the oscillator mechanism (4). This model predicts that these core clock genes also should influence the rhythmic expression of ''output'' genes important in regulating physiologic and biologic processes controlled by the circadian clock (5).Previous screens for such clock-controlled output genes have yielded varying estimates of their abundance and character in different organisms. An insertional reporter screen in the photosynthetic prokaryote Synechococcus suggested that most genes in this organism are transcribed in circadian fashion (6). Using microarray analysis, Harmer et al. identified 453 genes undergoing rhythmic expression under constant conditions in the plant Arabidopsis thaliana (7), representing Ϸ6% of the expressed genome. In Drosophila, analysis of 280 expressed sequence tags from the fly head revealed 20 diurnally varying transcripts, the majority of which were extremely rare, long messages of unclear physiologic function (8). The full extent of circadian gene expression is not known in any organism. The recent availability of an oligonucleotide-based microarray containing probes for nearly all known and predicted Drosophila genes allows estimation of the number of clock-controlled genes in the fly. Here we describe results of measuring circadian gene expression in control and period mutant flies in both light-dark (LD) and freerunning conditions. While this article was in preparation, three ot...