RESUMEN
Eukaryotic cells make many types of primary and processed RNAs that are found either in specific subcellular compartments or throughout the cells. A complete catalogue of these RNAs is not yet available and their characteristic subcellular localizations are also poorly understood. Because RNA represents the direct output of the genetic information encoded by genomes and a significant proportion of a cell's regulatory capabilities are focused on its synthesis, processing, transport, modification and translation, the generation of such a catalogue is crucial for understanding genome function. Here we report evidence that three-quarters of the human genome is capable of being transcribed, as well as observations about the range and levels of expression, localization, processing fates, regulatory regions and modifications of almost all currently annotated and thousands of previously unannotated RNAs. These observations, taken together, prompt a redefinition of the concept of a gene.
Asunto(s)
ADN/genética , Enciclopedias como Asunto , Genoma Humano/genética , Anotación de Secuencia Molecular , Secuencias Reguladoras de Ácidos Nucleicos/genética , Transcripción Genética/genética , Transcriptoma/genética , Alelos , Línea Celular , ADN Intergénico/genética , Elementos de Facilitación Genéticos , Exones/genética , Perfilación de la Expresión Génica , Genes/genética , Genómica , Humanos , Poliadenilación/genética , Isoformas de Proteínas/genética , ARN/biosíntesis , ARN/genética , Edición de ARN/genética , Empalme del ARN/genética , Secuencias Repetitivas de Ácidos Nucleicos/genética , Análisis de Secuencia de ARNRESUMEN
Drosophila melanogaster cell lines are important resources for cell biologists. Here, we catalog the expression of exons, genes, and unannotated transcriptional signals for 25 lines. Unannotated transcription is substantial (typically 19% of euchromatic signal). Conservatively, we identify 1405 novel transcribed regions; 684 of these appear to be new exons of neighboring, often distant, genes. Sixty-four percent of genes are expressed detectably in at least one line, but only 21% are detected in all lines. Each cell line expresses, on average, 5885 genes, including a common set of 3109. Expression levels vary over several orders of magnitude. Major signaling pathways are well represented: most differentiation pathways are "off" and survival/growth pathways "on." Roughly 50% of the genes expressed by each line are not part of the common set, and these show considerable individuality. Thirty-one percent are expressed at a higher level in at least one cell line than in any single developmental stage, suggesting that each line is enriched for genes characteristic of small sets of cells. Most remarkable is that imaginal disc-derived lines can generally be assigned, on the basis of expression, to small territories within developing discs. These mappings reveal unexpected stability of even fine-grained spatial determination. No two cell lines show identical transcription factor expression. We conclude that each line has retained features of an individual founder cell superimposed on a common "cell line" gene expression pattern.
Asunto(s)
Drosophila melanogaster/genética , Variación Genética , Transcripción Genética , Animales , Línea Celular , Análisis por Conglomerados , Exones , Femenino , Perfilación de la Expresión Génica , Masculino , Datos de Secuencia Molecular , Transducción de Señal/genética , Factores de Transcripción/genéticaRESUMEN
Large-scale sequencing projects have revealed an unexpected complexity in the origins, structures and functions of mammalian transcripts. Many loci are known to produce overlapping coding and noncoding RNAs with capped 5' ends that vary in size. Methods to identify the 5' ends of transcripts will facilitate the discovery of new promoters and 5' ends derived from secondary capping events. Such methods often require high input amounts of RNA not obtainable from highly refined samples such as tissue microdissections and subcellular fractions. Therefore, we developed nano-cap analysis of gene expression (nanoCAGE), a method that captures the 5' ends of transcripts from as little as 10 ng of total RNA, and CAGEscan, a mate-pair adaptation of nanoCAGE that captures the transcript 5' ends linked to a downstream region. Both of these methods allow further annotation-agnostic studies of the complex human transcriptome.