Metabolic labeling of RNA uncovers principles of RNA production and degradation dynamics in mammalian cells

• Published in: Nature biotechnology 2011-05, Vol.29 (5), p.436-442
• Main Authors: Rabani, Michal, Levin, Joshua Z, Fan, Lin, Adiconis, Xian, Raychowdhury, Raktima, Garber, Manuel, Gnirke, Andreas, Nusbaum, Chad, Hacohen, Nir, Friedman, Nir, Amit, Ido, Regev, Aviv
• Format: Article
• Language: English
• Subjects: 631/114/2397; 631/337/1645; 631/61/24; 631/61/32; Agriculture; Animals; Bioinformatics; Biological and medical sciences; Biomedical and Life Sciences; Biomedical Engineering/Biotechnology; Biomedicine; Biotechnology; Biotinylation - methods; Cells, Cultured; Cellular biology; Computational Biology; Degradation; Dendritic Cells - metabolism; Down-Regulation; Female; Fundamental and applied biological sciences. Psychology; Gene expression; Genetic Association Studies; Genomics; Immune system; Life Sciences; Lipopolysaccharides - metabolism; Mice; Mice, Inbred C57BL; Models, Molecular; Molecular and cellular biology; Molecular genetics; Physiological aspects; Proteolysis; Ribonucleic acid; RNA; RNA - genetics; RNA - metabolism; RNA Polymerase II - metabolism; RNA, Messenger - biosynthesis; RNA, Messenger - genetics; Sequence Analysis, RNA - methods; Transcription Factors - genetics; Transcription Factors - metabolism; Transcription, Genetic; Transcription. Transcription factor. Splicing. Rna processing; Up-Regulation
• ISSN: 1087-0156; 1546-1696
• DOI: 10.1038/nbt.1861

The relative contributions of RNA production, processing and degradation rates to cellular RNA levels are not well understood. Using pulsed metabolic RNA labeling of stimulated dendritic cells in conjunction with nCounter RNA quantification, RNA sequencing and computer modeling, Regev and colleagues unravel principles that determine RNA expression levels. Cellular RNA levels are determined by the interplay of RNA production, processing and degradation. However, because most studies of RNA regulation do not distinguish the separate contributions of these processes, little is known about how they are temporally integrated. Here we combine metabolic labeling of RNA at high temporal resolution with advanced RNA quantification and computational modeling to estimate RNA transcription and degradation rates during the response of mouse dendritic cells to lipopolysaccharide. We find that changes in transcription rates determine the majority of temporal changes in RNA levels, but that changes in degradation rates are important for shaping sharp 'peaked' responses. We used sequencing of the newly transcribed RNA population to estimate temporally constant RNA processing and degradation rates genome wide. Degradation rates vary significantly between genes and contribute to the observed differences in the dynamic response. Certain transcripts, including those encoding cytokines and transcription factors, mature faster. Our study provides a quantitative approach to study the integrative process of RNA regulation.