GSE25432
Temporal response of DCs to LPS stimulation: 4sU_sequencing
Regulation of RNA levels is critical for the response to external stimuli and determined through the interplay between RNA production, processing and degradation. Despite the centrality of these processes, most global studies of RNA regulation do not distinguish their separate contributions and relatively little is known about how they are temporally integrated. Here, we combine metabolic labeling of RNA with advanced RNA quantification assays and computational modeling to estimate RNA transcription and degradation during the response of immune dendritic cells (DCs) to pathogens, a critical and tightly regulated step in innate immunity. We find that transcription regulation plays a major role in shaping most temporal changes in RNA levels, but that changes in degradation rate are important for shaping sharp ‘peaked’ responses. We find that transcription changes precede corresponding RNA changes by a small lag (15-30 min), which is shorter for induced than for repressed genes. Massively parallel sequencing of the entire RNA population – including non-polyadenylated transcripts – allows us to estimate RNA processing, and identify specific groups of transcripts, mostly cytokines and transcription factors, undergoing enhanced mRNA maturation. This suggests an additional role for splicing in regulating mRNA maturation. Our method provides a new quantitative approach to study key steps in the integrative process of RNA regulation. Overall design: Sequencing of 4sU-labeled RNA taken from a 7 samples time-series (one sample every 1 hour) during the response of DCs to LPS stimulation. 4-thiouridine was added 45 minutes prior to sample collection. Data presented here for six timepoints: 0, 1, 3-6 hrs. 2hr timepoint not included.