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.

The recent advent of methods for high-throughput single-cell molecular profiling has catalyzed a growing sense in the scientific community that the time is ripe to complete the 150-year-old effort to identify all cell types in the human body. The Human Cell Atlas Project is an international collaborative effort that aims to define all human cell types in terms of distinctive molecular profiles (such as gene expression profiles) and to connect this information with classical cellular descriptions (such as location and morphology). An open comprehensive reference map of the molecular state of cells in healthy human tissues would propel the systematic study of physiological states, developmental trajectories, regulatory circuitry and interactions of cells, and also provide a framework for understanding cellular dysregulation in human disease. Here we describe the idea, its potential utility, early proofs-of-concept, and some design considerations for the Human Cell Atlas, including a commitment to open data, code, and community.

MicroRNAs (miRNAs) are short non-coding RNAs with key roles in cellular regulation. As part of the fifth edition of the Functional Annotation of Mammalian Genome (FANTOM5) project, we created an integrated expression atlas of miRNAs and their promoters by deep-sequencing 492 short RNA (sRNA) libraries, with matching Cap Analysis Gene Expression (CAGE) data, from 396 human and 47 mouse RNA samples. Promoters were identified for 1,357 human and 804 mouse miRNAs and showed strong sequence conservation between species. We also found that primary and mature miRNA expression levels were correlated, allowing us to use the primary miRNA measurements as a proxy for mature miRNA levels in a total of 1,829 human and 1,029 mouse CAGE libraries. We thus provide a broad atlas of miRNA expression and promoters in primary mammalian cells, establishing a foundation for detailed analysis of miRNA expression patterns and transcriptional control regions.

Kaposi sarcoma is considered a neoplasm of lymphatic endothelium infected with Kaposi sarcoma-associated herpesvirus. It is characterized by the expression of lymphatic lineage-specific genes by Kaposi sarcoma tumor cells. Here we show that infection of differentiated blood vascular endothelial cells with Kaposi sarcoma-associated herpesvirus leads to their lymphatic reprogramming; induction of ∼70% of the main lymphatic lineage-specific genes, including PROX1, a master regulator of lymphatic development; and downregulation of blood vascular genes.Kaposi sarcoma is the most frequently occurring malignant tumor in individuals infected with the human immunodeficiency (HIV) virus and also occurs in HIV-negative immunosuppressed individuals. Kaposi sarcoma mainly affects the skin and forms lesions of various types, including early inflammatory and patch stage lesions, and tumors with a predominant population of spindle cells. Infection with Kaposi sarcoma-associated herpesvirus (KSHV, also known as human herpesvirus-8) is essential for the formation of Kaposi sarcoma tumors 1,2 . Both latent viral genes, such as latency-associated nuclear antigen (LANA), and lytic viral genes, such as viral G-protein-coupled receptor, have been implicated in KSHV-mediated tumorigenesis 3 . In Kaposi sarcoma lesions, cells infected with KSHV characteristically appear spindle-shaped and are associated with slit-like spaces that sometimes contain red blood cells. Kaposi sarcoma is considered to be a neoplasm of KSHV-infected lymphatic endothelium, due to the morphological characteristics of the tumor cells and the expression of several lymphatic lineage-specific proteins, including VEGFR-3 and podoplanin [4][5][6][7] .The homeobox gene PROX1 is a master gene that controls lymphatic vessel development and differentiation 8 , and ectopic expression of PROX1 in differentiated blood vascular endothelial cells leads to lymphatic endothelial reprogramming of these cells 9,10 . Because KSHV can infect blood vascular endothelium, such as human umbilical vein endothelial cells, in vitro, we wondered whether KSHV infection might result in lymphatic reprogramming of blood vascular endothelium, potentially involving upregulation of PROX1.We first characterized the lineage-specific gene expression of cultured human lymphatic endothelial cells (LECs) versus blood vascular endothelial cells (BECs) 11 by Affymetrix HU133A gene arrays. We found that LECs, but not BECs, expressed PROX1, XLKD1 (encoding the hyaluronan receptor LYVE-1) and a number of other lymphatic lineage-specific genes ( Table 1). We then infected human dermal microvascular endothelial cells (HDMECs) with KSHV and carried out two independent transcriptional profiling studies 7 d later. Efficient KSHV infection was confirmed by high levels of expression of LANA mRNA in infected HDMECs. We found that 3-7% of infected HDMECs were ORF59-positive and 1.5-3% were K8.1 positive, in agreement with previous results [12][13][14] . This indicates that <10% of the cells were undergoing lytic reac...