The laboratory rat (Rattus norvegicus) is an indispensable tool in experimental medicine and drug development, having made inestimable contributions to human health. We report here the genome sequence of the Brown Norway (BN) rat strain. The sequence represents a high-quality 'draft' covering over 90% of the genome. The BN rat sequence is the third complete mammalian genome to be deciphered, and three-way comparisons with the human and mouse genomes resolve details of mammalian evolution. This first comprehensive analysis includes genes and proteins and their relation to human disease, repeated sequences, comparative genome-wide studies of mammalian orthologous chromosomal regions and rearrangement breakpoints, reconstruction of ancestral karyotypes and the events leading to existing species, rates of variation, and lineage-specific and lineage-independent evolutionary events such as expansion of gene families, orthology relations and protein evolution.

To examine the fundamental mechanisms governing neural differentiation, we analyzed the transcriptome changes that occur during the differentiation of hESCs into the neural lineage. Undifferentiated hESCs as well as cells at three stages of early neural differentiation-N1 (early initiation), N2 (neural progenitor), and N3 (early glial-like)-were analyzed using a combination of single read, paired-end read, and long read RNA sequencing. The results revealed enormous complexity in gene transcription and splicing dynamics during neural cell differentiation. We found previously unannotated transcripts and spliced isoforms specific for each stage of differentiation. Interestingly, splicing isoform diversity is highest in undifferentiated hESCs and decreases upon differentiation, a phenomenon we call isoform specialization. During neural differentiation, we observed differential expression of many types of genes, including those involved in key signaling pathways, and a large number of extracellular receptors exhibit stage-specific regulation. These results provide a valuable resource for studying neural differentiation and reveal insights into the mechanisms underlying in vitro neural differentiation of hESCs, such as neural fate specification, neural progenitor cell identity maintenance, and the transition from a predominantly neuronal state into one with increased gliogenic potential.RNA-Seq | splicing isoforms | unannotated transcripts | neuron | glial N eural commitment and subsequent differentiation is a complex process. Although the complexity of RNAs expressed in neural tissues is very high (1, 2), a comprehensive analysis of the genes and RNA isoforms that are expressed during the different stages of neural cell differentiation is largely lacking. Such information is expected to be important for understanding mechanisms of neural cell differentiation and ultimately providing therapeutic solutions for neural degenerative diseases, such as Parkinson's and Alzheimer's disease.Our current knowledge of the mechanisms involved in neural cell formation is derived mostly from studying neurogenesis in the developing embryos of animal models (3, 4). However, neurogenesis in animals is a complex process involving many different cell types that differentiate asynchronously. This heterogeneity, along with the relatively small number of cells that can be readily obtained, makes the analysis of the temporal differentiation of individual cell types extremely difficult. One solution is to analyze hESCs during in vitro differentiation to different stages of neural development, which can be performed using a relatively large numbers of cells (5-9). Analysis of the transcriptome in these cells is expected to provide insights into the mechanisms and pathways involved in early cell fate specification, such as the acquisition of neurogenic potential and the transition to gliogenic potential, which may ultimately be extremely useful for pharmacologic screening and neurodegenerative disease therapies.Many high-throughput methods have...

Alternative splicing (AS) dramatically expands the complexity of the mammalian brain transcriptome, but its atlas remains incomplete. Here we performed deep mRNA sequencing of mouse cortex to discover and characterize alternative exons with potential functional significance. Our analysis expands the list of AS events over 10-fold compared with previous annotations, demonstrating that 72% of multiexon genes express multiple splice variants in this single tissue. To evaluate functionality of the newly discovered AS events, we conducted comprehensive analyses on central nervous system (CNS) cell type-specific splicing, targets of tissue-or cell typespecific RNA binding proteins (RBPs), evolutionary selection pressure, and coupling of AS with nonsense-mediated decay (AS-NMD). We show that newly discovered events account for 23-42% of all cassette exons under tissue-or cell type-specific regulation. Furthermore, over 7,000 cassette exons are under evolutionary selection for regulated AS in mammals, 70% of which are new. Among these are 3,058 highly conserved cassette exons, including 1,014 NMD exons that may function directly to control gene expression levels. These NMD exons are particularly enriched in RBPs including splicing factors and interestingly also regulators for other steps of RNA metabolism. Unexpectedly, a second group of NMD exons reside in genes encoding chromatin regulators. Although the conservation of NMD exons in RBPs frequently extends into lower vertebrates, NMD exons in chromatin regulators are introduced later into the mammalian lineage, implying the emergence of a novel mechanism coupling AS and epigenetics. Our results highlight previously uncharacterized complexity and evolution in the mammalian brain transcriptome.new alternative exon | brain transcriptome | RNA-Seq | nonsense-mediated decay | chromatin regulator M olecular diversity derived from alternative splicing (AS) is believed to be critical for the creation of different cell types and tissues with distinct physiological properties and functions (1). This is particularly relevant to the central nervous system (CNS), which requires a large protein repertoire to generate its intricate and complex neural circuits (2). Therefore, a comprehensive catalog of AS events and identification of those with potential functional significance are important steps toward understanding the complexity of the nervous system.Over the past two decades, discovery and characterization of AS events using different technologies have provided important insights into the evolution and regulation of AS (3, 4). Earlier expressed sequence tag (EST)-based studies revealed the prevalence of AS in mammals (5). Investigation of these AS events, especially comparison of AS patterns in different species, led to an important observation that AS is rapidly evolving in mammals, with many alternative exons created after the split of primates and rodents (6). Evolutionarily recent exons in general have low level of inclusion and frequently result in frame shift and premature...

Toronto 2009 Data Release Workshop AuthorsOpen discussion of ideas and full disclosure of supporting facts provide the bedrock for scientific discourse and new developments. Traditionally, this has been formally accomplished through published papers, in which both the salient ideas and the supporting facts are combined in a single discrete 'package'. With the advent of methods for large-scale and high-throughput analyses, the generation and transmission of the underlying factual information -the data -are being transformed in an electronic process that involves submitting and retrieving information to and from scientific databases.

A critical problem in biology is understanding how cells choose between self-renewal and differentiation. To generate a comprehensive view of the mechanisms controlling early hematopoietic precursor self-renewal and differentiation, we used systems-based approaches and murine EML multipotential hematopoietic precursor cells as a primary model. EML cells give rise to a mixture of self-renewing Lin-SCA+CD34+ cells and partially differentiated non-renewing Lin-SCA-CD34− cells in a cell autonomous fashion. We identified and validated the HMG box protein TCF7 as a regulator in this self-renewal/differentiation switch that operates in the absence of autocrine Wnt signaling. We found that Tcf7 is the most down-regulated transcription factor when CD34+ cells switch into CD34− cells, using RNA–Seq. We subsequently identified the target genes bound by TCF7, using ChIP–Seq. We show that TCF7 and RUNX1 (AML1) bind to each other's promoter regions and that TCF7 is necessary for the production of the short isoforms, but not the long isoforms of RUNX1, suggesting that TCF7 and the short isoforms of RUNX1 function coordinately in regulation. Tcf7 knock-down experiments and Gene Set Enrichment Analyses suggest that TCF7 plays a dual role in promoting the expression of genes characteristic of self-renewing CD34+ cells while repressing genes activated in partially differentiated CD34− state. Finally a network of up-regulated transcription factors of CD34+ cells was constructed. Factors that control hematopoietic stem cell (HSC) establishment and development, cell growth, and multipotency were identified. These studies in EML cells demonstrate fundamental cell-intrinsic properties of the switch between self-renewal and differentiation, and yield valuable insights for manipulating HSCs and other differentiating systems.