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Key Points The mouse is the most widely used model organism to study human disease, but often mouse biology cannot be extrapolated to humans. A deep comparison of mouse and human physiology at the molecular level is essential for understanding under which circumstances the mouse can be a suitable model of human biology and for creating better mouse models. Advances in next-generation sequencing technologies fostered genome-wide annotation of functional DNA elements, enabling extensive comparison of the human and mouse genomes. At the transcriptional level, human and mouse gene expression profiles are conserved overall, although the degree of conservation varies depending on the tissues and the genes that are compared. Therefore, the question of whether the human and mouse transcriptomes cluster preferentially by tissue or organ or by species does not have an answer overall, and it depends specifically on the genes being considered. Conservation of expression is not a direct consequence of conservation in regulatory sequences, including promoters and enhancers. Although gene regulatory networks are preserved overall between human and mouse, transcription binding sites are often not conserved. Inter-individual genetic variation can affect human gene expression, but such variation cannot be modelled in inbred strains of laboratory mice because their genetic variation is small compared to the human population. An expansion of the current studies on the relationship between genetic variation and gene expression in outbred mice might provide helpful insights to understand the same relationship in humans. Emerging technologies — such single-cell genomics and single-cell spatial transcriptomics — and time series experiments will improve the annotation of human and mouse genomes, refine the current definitions of homologous cell types and homologous (molecular) phenotypes, and ultimately help scientists to identify which mouse models are the most appropriate to address a given biological question. Next-generation sequencing technologies have enabled the comprehensive characterization of human and mouse genomes, including at the transcriptional level. This article reviews the degree of conservation of human and mouse transcriptomes, along with the challenges of identifying when the mouse is a suitable model of human physiology. Cross-species comparisons of genomes, transcriptomes and gene regulation are now feasible at unprecedented resolution and throughput, enabling the comparison of human and mouse biology at the molecular level. Insights have been gained into the degree of conservation between human and mouse at the level of not only gene expression but also epigenetics and inter-individual variation. However, a number of limitations exist, including incomplete transcriptome characterization and difficulties in identifying orthologous phenotypes and cell types, which are beginning to be addressed by emerging technologies. Ultimately, these comparisons will help to identify the conditions under which the mouse is a suitable model of human physiology and disease, and optimize the use of animal models.

Deep sequencing of 'transcriptomes' — the collection of all RNA transcripts produced at a given time — from worms to humans reveals that some transcripts are composed of sequence segments that are not co-linear, with pieces of sequence coming from distant regions of DNA, even different chromosomes. Some of these 'chimaeric' transcripts are formed by genetic rearrangements, but others arise during post-transcriptional events. The ' trans -splicing' process in lower eukaryotes is well understood, but events in higher eukaryotes are not. The existence of such chimaeric RNAs has far-reaching implications for the potential information content of genomes and the way it is arranged.

Genes specifying long non-coding RNAs (lncRNAs) occupy a large fraction of the genomes of complex organisms. The term ‘lncRNAs’ encompasses RNA polymerase I (Pol I), Pol II and Pol III transcribed RNAs, and RNAs from processed introns. The various functions of lncRNAs and their many isoforms and interleaved relationships with other genes make lncRNA classification and annotation difficult. Most lncRNAs evolve more rapidly than protein-coding sequences, are cell type specific and regulate many aspects of cell differentiation and development and other physiological processes. Many lncRNAs associate with chromatin-modifying complexes, are transcribed from enhancers and nucleate phase separation of nuclear condensates and domains, indicating an intimate link between lncRNA expression and the spatial control of gene expression during development. lncRNAs also have important roles in the cytoplasm and beyond, including in the regulation of translation, metabolism and signalling. lncRNAs often have a modular structure and are rich in repeats, which are increasingly being shown to be relevant to their function. In this Consensus Statement, we address the definition and nomenclature of lncRNAs and their conservation, expression, phenotypic visibility, structure and functions. We also discuss research challenges and provide recommendations to advance the understanding of the roles of lncRNAs in development, cell biology and disease.This Consensus Statement addresses the definition, nomenclature and classification of long non-coding RNAs, and provides a shared viewpoint on their features and functions. The authors also discuss research challenges and provide recommendations to advance our understanding of long non-coding RNAs.

As a means to understand human neuropsychiatric disorders from human brain samples, we compared the transcription patterns and histological features of postmortem brain to fresh human neocortex isolated immediately following surgical removal. Compared to a number of neuropsychiatric disease-associated postmortem transcriptomes, the fresh human brain transcriptome had an entirely unique transcriptional pattern. To understand this difference, we measured genome-wide transcription as a function of time after fresh tissue removal to mimic the postmortem interval. Within a few hours, a selective reduction in the number of neuronal activity-dependent transcripts occurred with relative preservation of housekeeping genes commonly used as a reference for RNA normalization. Gene clustering indicated a rapid reduction in neuronal gene expression with a reciprocal time-dependent increase in astroglial and microglial gene expression that continued to increase for at least 24 h after tissue resection. Predicted transcriptional changes were confirmed histologically on the same tissue demonstrating that while neurons were degenerating, glial cells underwent an outgrowth of their processes. The rapid loss of neuronal genes and reciprocal expression of glial genes highlights highly dynamic transcriptional and cellular changes that occur during the postmortem interval. Understanding these time-dependent changes in gene expression in post mortem brain samples is critical for the interpretation of research studies on human brain disorders.

Although a small number of the vast array of animal long non-coding RNAs (lncRNAs) have known effects on cellular processes examined in vitro, the extent of their contributions to normal cell processes throughout development, differentiation and disease for the most part remains less clear. Phenotypes arising from deletion of an entire genomic locus cannot be unequivocally attributed either to the loss of the lncRNA per se or to the associated loss of other overlapping DNA regulatory elements. The distinction between cis- or trans-effects is also often problematic. We discuss the advantages and challenges associated with the current techniques for studying the in vivo function of lncRNAs in the light of different models of lncRNA molecular mechanism, and reflect on the design of experiments to mutate lncRNA loci. These considerations should assist in the further investigation of these transcriptional products of the genome.

Key Points In-depth analyses of the transcriptional outputs of eukaryotic genomes suggest that the information content of a genome is complex, and that this complexity manifests itself at two levels: the fraction of the genome that is devoted to encoding functional elements is higher than expected, and multiple functional elements can exist in a single region. The architecture of the eukaryotic transcriptome is clearly much more complex than could have been anticipated in terms of the number of nucleotides that are transcribed and the final arrangements of nucleotides that are present in mature processed RNA molecules. The complexity of genomic organization suggests that the currently accepted model, by which each region of DNA carries a single discrete function, must be re-evaluated, and an interleaved model for the arrangement of functional elements is more likely to represent the informational content of eukaryotic genomes. Despite the potential problems that are presented by use of the same genomic space for multiple purposes, the following advantages are brought by this complex genomic organization: an increase in protein-coding transcript diversity; a widespread adoption of RNA transcripts as regulatory agents; and a reliance on transcription as a regulatory process. On a global level, an interleaved genomic organization of functional elements seems to be preserved in different kingdoms, and the arrangement of specific overlapping functional elements is preserved among different species. This suggests that such a model does indeed provide advantages throughout evolution. Mutations at non-canonical sites, such as intronic regions that lie distal from splice sites, can affect fitness if they involve internal promoter regions, an exon of an overlapping transcript or a short RNA. Genome-wide analyses of transcriptional output in eukaryotes have revealed an unanticipated transcriptome complexity. These findings imply a complex, interleaved genomic organization, in which individual sequences carry multiple and overlapping informational content. The authors discuss the evidence for, and functional and evolutionary consequences of, this organization. Recent evidence of genome-wide transcription in several species indicates that the amount of transcription that occurs cannot be entirely accounted for by current sets of genome-wide annotations. Evidence indicates that most of both strands of the human genome might be transcribed, implying extensive overlap of transcriptional units and regulatory elements. These observations suggest that genomic architecture is not colinear, but is instead interleaved and modular, and that the same genomic sequences are multifunctional: that is, used for multiple independently regulated transcripts and as regulatory regions. What are the implications and consequences of such an interleaved genomic architecture in terms of increased information content, transcriptional complexity, evolution and disease states?

Significance To date, various studies have found similarities between humans and mice on a molecular level, and indeed, the murine model serves as an important experimental system for biomedical science. In this study of a broad number of tissues between humans and mice, high-throughput sequencing assays on the transcriptome and epigenome reveal that, in general, differences dominate similarities between the two species. These findings provide the basis for understanding the differences in phenotypes and responses to conditions in humans and mice. 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.

In parallel, whole chromosome tiling array interrogation of the RNA content of a variety of human tissues and cell lines revealed that, collectively, at least 93% of genomic bases are transcribed in one cell type or another [1],[10]-[13]. Since it is well established that highly expressed mRNAs dominate the non-ribosomal portion of the polyA+ transcriptome [7],[8],[10],[14]-[19], normalization approaches were used to reduce the quantity of highly expressed transcripts in these cDNA analyses [7],[8], and are implicit in tiling array approaches. [...]any estimate of the pervasiveness of transcription requires inclusion of all data sources, and less than exhaustive analyses can only provide lower bounds for transcriptional complexity.

Significant fractions of eukaryotic genomes give rise to RNA, much of which is unannotated and has reduced protein-coding potential. The genomic origins and the associations of human nuclear and cytosolic polyadenylated RNAs longer than 200 nucleotides (nt) and whole-cell RNAs less than 200 nt were investigated in this genome-wide study. Subcellular addresses for nucleotides present in detected RNAs were assigned, and their potential processing into short RNAs was investigated. Taken together, these observations suggest a novel role for some unannotated RNAs as primary transcripts for the production of short RNAs. Three potentially functional classes of RNAs have been identified, two of which are syntenically conserved and correlate with the expression state of protein-coding genes. These data support a highly interleaved organization of the human transcriptome.

I.L. was supported by a “Salvador Madariaga” fellowship from the Spanish Ministry of Science and Innovation. A.T.-A. and J.V. were supported by Spanish Ministry of Science and Innovation “Ramon y Cajal” contracts. M.V. was supported by a Consejo Superior de Investigaciones Científicas JAE Predoctoral research contract. This work was supported by Spanish Ministry of Science and Innovation Grants BIO2008-05284-C02-01 and ERA-NET Pathogenomics PIM2010EPA-00606.